p gingivalis w83 chromosomal dna  (Roche)


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    Structured Review

    Roche p gingivalis w83 chromosomal dna
    RT-PCR analysis of P. <t>gingivalis</t> <t>W83.</t> DNase treated total RNA was extracted from P. gingivalis W83 and isogenic mutants (FLL145, FLL146, FLL147 and FLL148) then subjected to RT-PCR. In Panel A , lanes 1, 2 and 3 shows expression of a 1.2 kb, 1.3 kb and 0.9 kb uvrA, pcrA and vimA fragments respectively in wild-type P. gingivalis W83; lane 4 shows no expression of the 1.2 kb uvrA fragment in the FLL145 mutant but shows expression of pcrA and vimA fragments as seen in lanes 5 and 6; in the FLL146 mutant, lanes 7 and 9 shows expression of uvrA and vimA fragments respectively but no expression of pcrA fragments is seen in lane 8; as a positive control, lanes 10, 11 and 12 shows expression of 16S ribosomal RNA. In Panel B , neither lanes 1 or 3 and 6 or 8 showed amplification of uvrA and pcrA fragments in FLL147 and FLL148 mutants respectively; lanes 2 and 7 showed amplification of PG1037 in FLL147 and FLL148 mutants respectively; lanes 4 and 9 shows amplification of vimA in FLL147 and FLL148 mutants respectively; lanes 5 and 10 shows amplification of 16S ribosomal RNA. In Panel C , lanes 1–5 shows that PG1037 is expressed in P. gingivalis W83 and all isogenic mutants; lanes 6–10 shows amplification of vimA in P. gingivalis W83 and all isogenic mutants.
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    Images

    1) Product Images from "Protective Role of the PG1036-PG1037-PG1038 Operon in Oxidative Stress in Porphyromonas gingivalis W83"

    Article Title: Protective Role of the PG1036-PG1037-PG1038 Operon in Oxidative Stress in Porphyromonas gingivalis W83

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0069645

    RT-PCR analysis of P. gingivalis W83. DNase treated total RNA was extracted from P. gingivalis W83 and isogenic mutants (FLL145, FLL146, FLL147 and FLL148) then subjected to RT-PCR. In Panel A , lanes 1, 2 and 3 shows expression of a 1.2 kb, 1.3 kb and 0.9 kb uvrA, pcrA and vimA fragments respectively in wild-type P. gingivalis W83; lane 4 shows no expression of the 1.2 kb uvrA fragment in the FLL145 mutant but shows expression of pcrA and vimA fragments as seen in lanes 5 and 6; in the FLL146 mutant, lanes 7 and 9 shows expression of uvrA and vimA fragments respectively but no expression of pcrA fragments is seen in lane 8; as a positive control, lanes 10, 11 and 12 shows expression of 16S ribosomal RNA. In Panel B , neither lanes 1 or 3 and 6 or 8 showed amplification of uvrA and pcrA fragments in FLL147 and FLL148 mutants respectively; lanes 2 and 7 showed amplification of PG1037 in FLL147 and FLL148 mutants respectively; lanes 4 and 9 shows amplification of vimA in FLL147 and FLL148 mutants respectively; lanes 5 and 10 shows amplification of 16S ribosomal RNA. In Panel C , lanes 1–5 shows that PG1037 is expressed in P. gingivalis W83 and all isogenic mutants; lanes 6–10 shows amplification of vimA in P. gingivalis W83 and all isogenic mutants.
    Figure Legend Snippet: RT-PCR analysis of P. gingivalis W83. DNase treated total RNA was extracted from P. gingivalis W83 and isogenic mutants (FLL145, FLL146, FLL147 and FLL148) then subjected to RT-PCR. In Panel A , lanes 1, 2 and 3 shows expression of a 1.2 kb, 1.3 kb and 0.9 kb uvrA, pcrA and vimA fragments respectively in wild-type P. gingivalis W83; lane 4 shows no expression of the 1.2 kb uvrA fragment in the FLL145 mutant but shows expression of pcrA and vimA fragments as seen in lanes 5 and 6; in the FLL146 mutant, lanes 7 and 9 shows expression of uvrA and vimA fragments respectively but no expression of pcrA fragments is seen in lane 8; as a positive control, lanes 10, 11 and 12 shows expression of 16S ribosomal RNA. In Panel B , neither lanes 1 or 3 and 6 or 8 showed amplification of uvrA and pcrA fragments in FLL147 and FLL148 mutants respectively; lanes 2 and 7 showed amplification of PG1037 in FLL147 and FLL148 mutants respectively; lanes 4 and 9 shows amplification of vimA in FLL147 and FLL148 mutants respectively; lanes 5 and 10 shows amplification of 16S ribosomal RNA. In Panel C , lanes 1–5 shows that PG1037 is expressed in P. gingivalis W83 and all isogenic mutants; lanes 6–10 shows amplification of vimA in P. gingivalis W83 and all isogenic mutants.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, Mutagenesis, Positive Control, Amplification

    UV sensitivity of P. gingivalis mutants. P. gingivalis strains W83, FLL144, FLL145 and FLL146 were grown to mid log phase (OD 600 of 0.6) spread on BHI plates then subjected to irradiation at increasing doses (0 µJ, 500 µJ and 1000 µJ) of UV in a Stratalinker 2400 (Stratagene, La Jolla, CA). **P≤0.01, *P≤0.05.
    Figure Legend Snippet: UV sensitivity of P. gingivalis mutants. P. gingivalis strains W83, FLL144, FLL145 and FLL146 were grown to mid log phase (OD 600 of 0.6) spread on BHI plates then subjected to irradiation at increasing doses (0 µJ, 500 µJ and 1000 µJ) of UV in a Stratalinker 2400 (Stratagene, La Jolla, CA). **P≤0.01, *P≤0.05.

    Techniques Used: Irradiation

    8-oxoG removal activities of cell extracts from P. gingivalis strains W83 and isogenic mutants. [γ- 32 P]-ATP-5′-end-labeled oligonucleotides (O1 and O2) were incubated with P. gingivalis extracts for 10 min, electrophoresed and visualized. Lane 1 contained O1; Lane 2 contained O2; Lane 3 contained O1 and Fpg enzyme; Lane 4 contained O2 and Fpg enzyme; Lane 5 contained O1 and P. gingivalis W83 extract; Lane 6 contained O2 and P. gingivalis W83 extract; Lane 7 contained O1 and P. gingivalis FLL145 extract; Lane 8 contained O2 and P. gingivalis FLL145 extract; Lane 9 contained O1 and P. gingivalis FLL146 extract; Lane 10 contained O2 and P. gingivalis FLL146 extract; Lane 11 contained O1 and P. gingivalis FLL147 extract; Lane 12 contained O2 and P. gingivalis FLL147 extract; Lane 13 contained O1 and P. gingivalis FLL148 extract and Lane 14 contained O2 and P. gingivalis FLL148 extract.
    Figure Legend Snippet: 8-oxoG removal activities of cell extracts from P. gingivalis strains W83 and isogenic mutants. [γ- 32 P]-ATP-5′-end-labeled oligonucleotides (O1 and O2) were incubated with P. gingivalis extracts for 10 min, electrophoresed and visualized. Lane 1 contained O1; Lane 2 contained O2; Lane 3 contained O1 and Fpg enzyme; Lane 4 contained O2 and Fpg enzyme; Lane 5 contained O1 and P. gingivalis W83 extract; Lane 6 contained O2 and P. gingivalis W83 extract; Lane 7 contained O1 and P. gingivalis FLL145 extract; Lane 8 contained O2 and P. gingivalis FLL145 extract; Lane 9 contained O1 and P. gingivalis FLL146 extract; Lane 10 contained O2 and P. gingivalis FLL146 extract; Lane 11 contained O1 and P. gingivalis FLL147 extract; Lane 12 contained O2 and P. gingivalis FLL147 extract; Lane 13 contained O1 and P. gingivalis FLL148 extract and Lane 14 contained O2 and P. gingivalis FLL148 extract.

    Techniques Used: Labeling, Incubation

    RT-PCR analysis of P. gingivalis W83. DNase treated total RNA extracted from P. gingivalis W83 and subjected to RT-PCR. Lane 1, amplification of 4.3 kb uvrA-PG1037 fragment; Lane 2, amplification of 3.7 kb PG1037-pcrA fragment; Lanes 3, 4 and 5, intragenic specific primers for uvrA , PG1037 and pcrA genes amplified a 1.2 kb, 1.0 kb and 1.3 kb fragments respectively; Lane 6, amplification of 0.7 kb 16S fragment.
    Figure Legend Snippet: RT-PCR analysis of P. gingivalis W83. DNase treated total RNA extracted from P. gingivalis W83 and subjected to RT-PCR. Lane 1, amplification of 4.3 kb uvrA-PG1037 fragment; Lane 2, amplification of 3.7 kb PG1037-pcrA fragment; Lanes 3, 4 and 5, intragenic specific primers for uvrA , PG1037 and pcrA genes amplified a 1.2 kb, 1.0 kb and 1.3 kb fragments respectively; Lane 6, amplification of 0.7 kb 16S fragment.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Amplification

    2) Product Images from "Linkage Group Selection: Towards Identifying Genes Controlling Strain Specific Protective Immunity in Malaria"

    Article Title: Linkage Group Selection: Towards Identifying Genes Controlling Strain Specific Protective Immunity in Malaria

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0000857

    The Comparative Intensities of 92 AFLP markers of Plasmodium chabaudi chabaudi strain AJ from the progeny of a genetic cross between P. c. chabaudi strains CB-pyr10 and AJ following selection in mice immunised strain AJ (see text). AJ-specific markers (indicated by black diamonds) were located on a P. c. chabaudi genetic linkage map, generated from a genetic cross between AS and AJ [34] . Numbers after letter ‘C’ and ‘g’ represent P. c. chabaudi chromosome numbers and P. c. chabaudi unassigned linkage groups, respectively, in the genetic linkage map [34] . Of the six AJ markers which were most reduced under AJ-specific immune selection (see Table 2 ), five (indicated by asterisks) could be located to P. c. chabaudi chromosome 8, forming a selection valley with the P. c. chabaudi msp -1 gene at its lowest point (see text). RTQ-PCR values for the proportions of the AJ-immune selected cross progeny carrying the AJ alleles of the Merozoite Surface Protein-1 ( msp -1) are indicated by the red triangle and Apical Membrane Antigen-1 ( ama -1) by the green triangle in the AJ-immune selected cross progeny. The red line indicates Comparative Intensity of 50%.
    Figure Legend Snippet: The Comparative Intensities of 92 AFLP markers of Plasmodium chabaudi chabaudi strain AJ from the progeny of a genetic cross between P. c. chabaudi strains CB-pyr10 and AJ following selection in mice immunised strain AJ (see text). AJ-specific markers (indicated by black diamonds) were located on a P. c. chabaudi genetic linkage map, generated from a genetic cross between AS and AJ [34] . Numbers after letter ‘C’ and ‘g’ represent P. c. chabaudi chromosome numbers and P. c. chabaudi unassigned linkage groups, respectively, in the genetic linkage map [34] . Of the six AJ markers which were most reduced under AJ-specific immune selection (see Table 2 ), five (indicated by asterisks) could be located to P. c. chabaudi chromosome 8, forming a selection valley with the P. c. chabaudi msp -1 gene at its lowest point (see text). RTQ-PCR values for the proportions of the AJ-immune selected cross progeny carrying the AJ alleles of the Merozoite Surface Protein-1 ( msp -1) are indicated by the red triangle and Apical Membrane Antigen-1 ( ama -1) by the green triangle in the AJ-immune selected cross progeny. The red line indicates Comparative Intensity of 50%.

    Techniques Used: Selection, Mouse Assay, Generated, Polymerase Chain Reaction

    3) Product Images from "Regulation of the Expression of the Vibrio parahaemolyticus peuA Gene Encoding an Alternative Ferric Enterobactin Receptor"

    Article Title: Regulation of the Expression of the Vibrio parahaemolyticus peuA Gene Encoding an Alternative Ferric Enterobactin Receptor

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0105749

    Genetic map and operon structure of VPA0148 – VPA0156 locus. (A) Genetic map of the peuA gene and the flanking genes. Thick arrows indicate genes and their orientations. The –35 and –10 promoter elements and putative Fur box sequences in the promoter regions of peuR ( VPA0148 ) and peuA ( VPA0150 ) are indicated. The transcription start sites for peuR (+1) and peuA (+1 and +39) are indicated by right-angled arrows. The putative terminator signal between the peuS and peuA genes, the predicted RBS for the peuA gene, the start codons for peuR and peuA genes, and the stop codon for peuS are also indicated. The amino acid sequence consistent with the N-terminal sequence determined for the iron-repressible OMR induced in LB-Tris/+EDDA and LB-Tris/+EDDA/+Ent media at pH 8.0 (see Figure 3 ) is indicated by a double underline. (B) Schematic representation of mRNAs transcribed from the VPA0148 - VPA0156 genes and the primer pairs used for RT-PCR. For preparation of cDNAs by RT, VPpeuS-R and VP0156-R were used. (C) RT-PCR analysis of RT-PCR products. +RT and –RT, RT-PCR was performed with and without reverse transcriptase, respectively. M, 100-bp DNA ladder.
    Figure Legend Snippet: Genetic map and operon structure of VPA0148 – VPA0156 locus. (A) Genetic map of the peuA gene and the flanking genes. Thick arrows indicate genes and their orientations. The –35 and –10 promoter elements and putative Fur box sequences in the promoter regions of peuR ( VPA0148 ) and peuA ( VPA0150 ) are indicated. The transcription start sites for peuR (+1) and peuA (+1 and +39) are indicated by right-angled arrows. The putative terminator signal between the peuS and peuA genes, the predicted RBS for the peuA gene, the start codons for peuR and peuA genes, and the stop codon for peuS are also indicated. The amino acid sequence consistent with the N-terminal sequence determined for the iron-repressible OMR induced in LB-Tris/+EDDA and LB-Tris/+EDDA/+Ent media at pH 8.0 (see Figure 3 ) is indicated by a double underline. (B) Schematic representation of mRNAs transcribed from the VPA0148 - VPA0156 genes and the primer pairs used for RT-PCR. For preparation of cDNAs by RT, VPpeuS-R and VP0156-R were used. (C) RT-PCR analysis of RT-PCR products. +RT and –RT, RT-PCR was performed with and without reverse transcriptase, respectively. M, 100-bp DNA ladder.

    Techniques Used: Sequencing, Reverse Transcription Polymerase Chain Reaction

    Primer extension analyses of total RNA from VPD54 or VPD102 to determine the transcription start sites of peuR (A) and peuA (B and C). Total RNAs were isolated from VPD54 ( vctA - and irgA -deficient mutant derived from VPD5) and VPD102 ( peuRS -deficient mutant derived from VPD54) grown at pH 7.0 and 8.0 in LB-Tris, LB-Tris/+EDDA, and LB-Tris/+EDDA/+Ent media. The amounts of total RNA and primers used for reverse transcription were as follows: (A) 10 µg VppeuR-PE, (B) 10 µg VppeuA-PE, and (C) 150 µg VppeuA-PE. The same primers used for primer extension analysis were used to generate the sequence ladders (A, C, G, T). The transcription start sites and putative Fur boxes are indicated at the top of panels A and B (also see Figure 3A ).
    Figure Legend Snippet: Primer extension analyses of total RNA from VPD54 or VPD102 to determine the transcription start sites of peuR (A) and peuA (B and C). Total RNAs were isolated from VPD54 ( vctA - and irgA -deficient mutant derived from VPD5) and VPD102 ( peuRS -deficient mutant derived from VPD54) grown at pH 7.0 and 8.0 in LB-Tris, LB-Tris/+EDDA, and LB-Tris/+EDDA/+Ent media. The amounts of total RNA and primers used for reverse transcription were as follows: (A) 10 µg VppeuR-PE, (B) 10 µg VppeuA-PE, and (C) 150 µg VppeuA-PE. The same primers used for primer extension analysis were used to generate the sequence ladders (A, C, G, T). The transcription start sites and putative Fur boxes are indicated at the top of panels A and B (also see Figure 3A ).

    Techniques Used: Isolation, Mutagenesis, Derivative Assay, Sequencing

    In vitro translation of peuA mRNA. (A) In vitro translation analysis of the +1 and +39 peuA transcripts labeled with the FLAG tag. The +1- peuA ′- flag RNA and +39- peuA ′- flag RNA were first synthesized by in vitro transcription, as described in the MATERIALS AND METHODS, and a mixture containing either the +1- peuA ′- flag RNA (30 pmol)/ fur - flag RNA (3 pmol) or the +39- peuA ′- flag RNA (30 pmol)/ fur - flag RNA (3 pmol) as the template was subjected to in vitro translation. The FLAG-fused proteins translated were separated on 15% SDS-polyacrylamide gels, and were detected by western blotting using anti-FLAG IgG. (B) Confirmation of the presence of +1- peuA ′- flag RNA and +39- peuA ′- flag RNA in the reaction mixture for in vitro translation. These RNA fragments were detected in the reaction mixture by northern blotting using a DIG-labeled peuA probe.
    Figure Legend Snippet: In vitro translation of peuA mRNA. (A) In vitro translation analysis of the +1 and +39 peuA transcripts labeled with the FLAG tag. The +1- peuA ′- flag RNA and +39- peuA ′- flag RNA were first synthesized by in vitro transcription, as described in the MATERIALS AND METHODS, and a mixture containing either the +1- peuA ′- flag RNA (30 pmol)/ fur - flag RNA (3 pmol) or the +39- peuA ′- flag RNA (30 pmol)/ fur - flag RNA (3 pmol) as the template was subjected to in vitro translation. The FLAG-fused proteins translated were separated on 15% SDS-polyacrylamide gels, and were detected by western blotting using anti-FLAG IgG. (B) Confirmation of the presence of +1- peuA ′- flag RNA and +39- peuA ′- flag RNA in the reaction mixture for in vitro translation. These RNA fragments were detected in the reaction mixture by northern blotting using a DIG-labeled peuA probe.

    Techniques Used: In Vitro, Labeling, FLAG-tag, Synthesized, Western Blot, Northern Blot

    4) Product Images from "Comparison of PRRSV Nucleic Acid and Antibody Detection in Pen-Based Oral Fluid and Individual Serum Samples in Three Different Age Categories of Post-Weaning Pigs from Endemically Infected Farms"

    Article Title: Comparison of PRRSV Nucleic Acid and Antibody Detection in Pen-Based Oral Fluid and Individual Serum Samples in Three Different Age Categories of Post-Weaning Pigs from Endemically Infected Farms

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0166300

    Overview of PRRSV detection by qRT-PCR in serum and oral fluid collected on seven endemically PRRSV infected farms (F1-F7) and one PRRSV free farm (F9). Full dots indicate the percentage of PRRSV positive pigs in serum per pen. If the full dot is surrounded by an open circle, it indicates that the corresponding pen based oral fluid sample was positive in qRT-PCR.
    Figure Legend Snippet: Overview of PRRSV detection by qRT-PCR in serum and oral fluid collected on seven endemically PRRSV infected farms (F1-F7) and one PRRSV free farm (F9). Full dots indicate the percentage of PRRSV positive pigs in serum per pen. If the full dot is surrounded by an open circle, it indicates that the corresponding pen based oral fluid sample was positive in qRT-PCR.

    Techniques Used: Quantitative RT-PCR, Infection

    Probability to detect PRRSV in a pen based oral fluid sample by qRT-PCR dependent on the percentage of PRRSV positive pigs in serum per pen.
    Figure Legend Snippet: Probability to detect PRRSV in a pen based oral fluid sample by qRT-PCR dependent on the percentage of PRRSV positive pigs in serum per pen.

    Techniques Used: Quantitative RT-PCR

    5) Product Images from "Detection of RUNX2 gene expression in cumulus cells in women undergoing controlled ovarian stimulation"

    Article Title: Detection of RUNX2 gene expression in cumulus cells in women undergoing controlled ovarian stimulation

    Journal: Reproductive Biology and Endocrinology : RB & E

    doi: 10.1186/1477-7827-10-99

    Distribution of RUNX2 gene expression (A) on patient’s population and statistically significant correlations between ovulation induction factors, such as number of follicles, oocytes, fertilized oocytes (B) and hormone serum levels for E2 (C) and LH (D) respectively. RUNX2 gene expression was determined by real-time PCR for 41 patients and categorized into two groups of patients, with expression and without expression. Expression ratios (RUNX2 copies/G6PDH copies) were used for the evaluation of RUNX2 gene expression.
    Figure Legend Snippet: Distribution of RUNX2 gene expression (A) on patient’s population and statistically significant correlations between ovulation induction factors, such as number of follicles, oocytes, fertilized oocytes (B) and hormone serum levels for E2 (C) and LH (D) respectively. RUNX2 gene expression was determined by real-time PCR for 41 patients and categorized into two groups of patients, with expression and without expression. Expression ratios (RUNX2 copies/G6PDH copies) were used for the evaluation of RUNX2 gene expression.

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction

    6) Product Images from "Effect of Intragenic Rearrangement and Changes in the 3? Consensus Sequence on NSP1 Expression and Rotavirus Replication"

    Article Title: Effect of Intragenic Rearrangement and Changes in the 3? Consensus Sequence on NSP1 Expression and Rotavirus Replication

    Journal: Journal of Virology

    doi: 10.1128/JVI.75.5.2076-2086.2001

    Effect of the UGACC→UGAACC mutation on protein expression in infected cells. The chimeric reporter RNAs g6-Fluc-UGACC and -UGAACC and glo/g6-Fluc-UGACC and -UGAACC were separately cotransfected with nv-Rluc into SA11-infected MA104 cells at 1 h p.i. At 9 h p.i., the levels of firefly and Renilla luciferases per milligram of cell lysate were determined and the expression of firefly luciferase was normalized to the expression of Renilla luciferase. To ease the comparison of values, the expression of firefly luciferase for the g6-Fluc-UGACC and glo/g6-Fluc-UGACC RNAs was set to 100%.
    Figure Legend Snippet: Effect of the UGACC→UGAACC mutation on protein expression in infected cells. The chimeric reporter RNAs g6-Fluc-UGACC and -UGAACC and glo/g6-Fluc-UGACC and -UGAACC were separately cotransfected with nv-Rluc into SA11-infected MA104 cells at 1 h p.i. At 9 h p.i., the levels of firefly and Renilla luciferases per milligram of cell lysate were determined and the expression of firefly luciferase was normalized to the expression of Renilla luciferase. To ease the comparison of values, the expression of firefly luciferase for the g6-Fluc-UGACC and glo/g6-Fluc-UGACC RNAs was set to 100%.

    Techniques Used: Mutagenesis, Expressing, Infection, Luciferase

    7) Product Images from "Detection of Severe Acute Respiratory Syndrome Coronavirus in Blood of Infected Patients"

    Article Title: Detection of Severe Acute Respiratory Syndrome Coronavirus in Blood of Infected Patients

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.42.1.347-350.2004

    Detection of SARS CoV from 18 subjects by real-time quantitative PCR. Data for RNA from 18 subjects are shown. The viral load is indicated as number of copies per ml of blood. Subject 1, 2,020 copies; subject 2, 9,100 copies; subject 3, 3,120 copies; subject 4, 9,600 copies. A sample would be considered positive if it generated a typical amplification curve within the 45 cycles. Negative signals from subjects 5 to 18 and the nontemplate control (water) are shown. The inset graph shows melting-curve analysis of the PCR products from the 18 subjects. The x axis denotes the temperature ( o C), and the y axis denotes the fluorescence intensity (F2) over the background level.
    Figure Legend Snippet: Detection of SARS CoV from 18 subjects by real-time quantitative PCR. Data for RNA from 18 subjects are shown. The viral load is indicated as number of copies per ml of blood. Subject 1, 2,020 copies; subject 2, 9,100 copies; subject 3, 3,120 copies; subject 4, 9,600 copies. A sample would be considered positive if it generated a typical amplification curve within the 45 cycles. Negative signals from subjects 5 to 18 and the nontemplate control (water) are shown. The inset graph shows melting-curve analysis of the PCR products from the 18 subjects. The x axis denotes the temperature ( o C), and the y axis denotes the fluorescence intensity (F2) over the background level.

    Techniques Used: Real-time Polymerase Chain Reaction, Generated, Amplification, Polymerase Chain Reaction, Fluorescence

    Detection of SARS CoV by real-time quantitative PCR. (A) Amplification of single-stranded RNA standards (indicated as a to e) and RNA extracted from sputum, stool, and blood spiked with virus grown in Vero E6 cells. The x axis denotes the cycle number of the quantitative PCR assay, and the y axis denotes fluorescence intensity (F2) over the background level. The nontemplate control (water) is indicated. The viral load is indicated as the number of copies per reaction: spiked sputum, 5 × 10 3 ; spiked stool, 4 × 10 3 ; spiked blood, 1 × 10 3 . RNA standards were as follows: (a) 1 × 10 6 copies per reaction, (b) 9.5 × 10 4 copies per reaction, (c) 8.7 × 10 3 copies per reaction, (d) 1.1 × 10 3 copies per reaction, and (e) 8.5 × 10 copies per reaction. The inset graph represents melting-curve analysis of the PCR products. Signals from RNA standards (a to e), spiked samples, normal samples, and nontemplate control (water) are shown. The x axis denotes the temperature ( o C), and the y axis denotes the fluorescence intensity (F2) over the background level. (B) Detection of the internal control in fluorometer channel F3 in parallel with the simultaneous amplification of the RNA standards (a to e), spiked samples, and nontemplate control is shown.
    Figure Legend Snippet: Detection of SARS CoV by real-time quantitative PCR. (A) Amplification of single-stranded RNA standards (indicated as a to e) and RNA extracted from sputum, stool, and blood spiked with virus grown in Vero E6 cells. The x axis denotes the cycle number of the quantitative PCR assay, and the y axis denotes fluorescence intensity (F2) over the background level. The nontemplate control (water) is indicated. The viral load is indicated as the number of copies per reaction: spiked sputum, 5 × 10 3 ; spiked stool, 4 × 10 3 ; spiked blood, 1 × 10 3 . RNA standards were as follows: (a) 1 × 10 6 copies per reaction, (b) 9.5 × 10 4 copies per reaction, (c) 8.7 × 10 3 copies per reaction, (d) 1.1 × 10 3 copies per reaction, and (e) 8.5 × 10 copies per reaction. The inset graph represents melting-curve analysis of the PCR products. Signals from RNA standards (a to e), spiked samples, normal samples, and nontemplate control (water) are shown. The x axis denotes the temperature ( o C), and the y axis denotes the fluorescence intensity (F2) over the background level. (B) Detection of the internal control in fluorometer channel F3 in parallel with the simultaneous amplification of the RNA standards (a to e), spiked samples, and nontemplate control is shown.

    Techniques Used: Real-time Polymerase Chain Reaction, Amplification, Fluorescence, Polymerase Chain Reaction

    8) Product Images from "Linkage Group Selection: Towards Identifying Genes Controlling Strain Specific Protective Immunity in Malaria"

    Article Title: Linkage Group Selection: Towards Identifying Genes Controlling Strain Specific Protective Immunity in Malaria

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0000857

    The Comparative Intensities of 92 AFLP markers of Plasmodium chabaudi chabaudi strain AJ from the progeny of a genetic cross between P. c. chabaudi strains CB-pyr10 and AJ following selection in mice immunised strain AJ (see text). AJ-specific markers (indicated by black diamonds) were located on a P. c. chabaudi genetic linkage map, generated from a genetic cross between AS and AJ [34] . Numbers after letter ‘C’ and ‘g’ represent P. c. chabaudi chromosome numbers and P. c. chabaudi unassigned linkage groups, respectively, in the genetic linkage map [34] . Of the six AJ markers which were most reduced under AJ-specific immune selection (see Table 2 ), five (indicated by asterisks) could be located to P. c. chabaudi chromosome 8, forming a selection valley with the P. c. chabaudi msp -1 gene at its lowest point (see text). RTQ-PCR values for the proportions of the AJ-immune selected cross progeny carrying the AJ alleles of the Merozoite Surface Protein-1 ( msp -1) are indicated by the red triangle and Apical Membrane Antigen-1 ( ama -1) by the green triangle in the AJ-immune selected cross progeny. The red line indicates Comparative Intensity of 50%.
    Figure Legend Snippet: The Comparative Intensities of 92 AFLP markers of Plasmodium chabaudi chabaudi strain AJ from the progeny of a genetic cross between P. c. chabaudi strains CB-pyr10 and AJ following selection in mice immunised strain AJ (see text). AJ-specific markers (indicated by black diamonds) were located on a P. c. chabaudi genetic linkage map, generated from a genetic cross between AS and AJ [34] . Numbers after letter ‘C’ and ‘g’ represent P. c. chabaudi chromosome numbers and P. c. chabaudi unassigned linkage groups, respectively, in the genetic linkage map [34] . Of the six AJ markers which were most reduced under AJ-specific immune selection (see Table 2 ), five (indicated by asterisks) could be located to P. c. chabaudi chromosome 8, forming a selection valley with the P. c. chabaudi msp -1 gene at its lowest point (see text). RTQ-PCR values for the proportions of the AJ-immune selected cross progeny carrying the AJ alleles of the Merozoite Surface Protein-1 ( msp -1) are indicated by the red triangle and Apical Membrane Antigen-1 ( ama -1) by the green triangle in the AJ-immune selected cross progeny. The red line indicates Comparative Intensity of 50%.

    Techniques Used: Selection, Mouse Assay, Generated, Polymerase Chain Reaction

    Mixed strain infections of Plasmodium chabaudi chabaudi CB and AJ in mice pre-immunised with either strain, or in a non-immune batch mate. (A) shows total parasitaemias during the course of mixed strain infections in immunised and non-immune mice as measured by microscopic examination of thin blood smears stained with Giemsa's stain: non-immune mouse (dotted line with open circles), two CB-immunised mice (dashed lines with filled symbols), two AJ-immunised mice (unbroken lines with filled symbols). Strain specific parasitaemias (AJ in pink; CB in green) are shown (B) in the non-immune mouse, (C) in the two CB-immunised mice and (D) in the two AJ-immunised mice. The strain specific parasitaemias in the mixed strain infections were calculated from the total parasitaemias measured on thin blood smears stained with Giemsa's stain and from the proportions of CB and AJ parasites in the mixtures as determined using strain specific RTQ-PCR (see text). Squares and triangles represent mouse 1 and 2, respectively, in the CB and AJ immunised mice in Fig. 1A, C and D.
    Figure Legend Snippet: Mixed strain infections of Plasmodium chabaudi chabaudi CB and AJ in mice pre-immunised with either strain, or in a non-immune batch mate. (A) shows total parasitaemias during the course of mixed strain infections in immunised and non-immune mice as measured by microscopic examination of thin blood smears stained with Giemsa's stain: non-immune mouse (dotted line with open circles), two CB-immunised mice (dashed lines with filled symbols), two AJ-immunised mice (unbroken lines with filled symbols). Strain specific parasitaemias (AJ in pink; CB in green) are shown (B) in the non-immune mouse, (C) in the two CB-immunised mice and (D) in the two AJ-immunised mice. The strain specific parasitaemias in the mixed strain infections were calculated from the total parasitaemias measured on thin blood smears stained with Giemsa's stain and from the proportions of CB and AJ parasites in the mixtures as determined using strain specific RTQ-PCR (see text). Squares and triangles represent mouse 1 and 2, respectively, in the CB and AJ immunised mice in Fig. 1A, C and D.

    Techniques Used: Mouse Assay, Staining, Polymerase Chain Reaction

    9) Product Images from "Identification of HIV Superinfection in Seroconcordant Couples in Rakai, Uganda, by Use of Next-Generation Deep Sequencing ▿Identification of HIV Superinfection in Seroconcordant Couples in Rakai, Uganda, by Use of Next-Generation Deep Sequencing ▿ †"

    Article Title: Identification of HIV Superinfection in Seroconcordant Couples in Rakai, Uganda, by Use of Next-Generation Deep Sequencing ▿Identification of HIV Superinfection in Seroconcordant Couples in Rakai, Uganda, by Use of Next-Generation Deep Sequencing ▿ †

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.00804-11

    Inter- and intrasubtype detection of the p24 region by NGS. Neighbor-joining trees of p24 next-generation consensus sequences (≥10 identical reads) of intersubtype mixtures of A 2 and D 1 (red) at 95:5 (A) and 99:1 (B) ratios are shown. Intrasubtype mixtures of A 2 and A 1 (C; blue) and D 3 and D 4 (D; green) at the ratio of 95:5 are shown. The trees were constructed with a selection of subtype reference sequences and random sequences from individuals in Rakai shown in black. Brackets demonstrate the source of different clades within the merged trees. Bootstrap values higher than 80% are shown (1,000 replicates).
    Figure Legend Snippet: Inter- and intrasubtype detection of the p24 region by NGS. Neighbor-joining trees of p24 next-generation consensus sequences (≥10 identical reads) of intersubtype mixtures of A 2 and D 1 (red) at 95:5 (A) and 99:1 (B) ratios are shown. Intrasubtype mixtures of A 2 and A 1 (C; blue) and D 3 and D 4 (D; green) at the ratio of 95:5 are shown. The trees were constructed with a selection of subtype reference sequences and random sequences from individuals in Rakai shown in black. Brackets demonstrate the source of different clades within the merged trees. Bootstrap values higher than 80% are shown (1,000 replicates).

    Techniques Used: Next-Generation Sequencing, Construct, Selection

    Mixture analysis of intersubtype detection by NGS. Neighbor-joining trees of p24 next-generation consensus sequences (≥10 identical reads) of control samples of A 2 (A; blue), subtypes D 1 (B; green), a mixture of A 2 and D 1 at a 50:50 ratio (C; red), and a merged tree of all three sample runs (D) are shown. The trees were constructed with a selection of subtype reference sequences and random sequences from individuals in Rakai shown in black. Brackets demonstrate the source of different clades within the merged trees. Bootstrap values higher than 80% are shown for nonmerged trees (1,000 replicates).
    Figure Legend Snippet: Mixture analysis of intersubtype detection by NGS. Neighbor-joining trees of p24 next-generation consensus sequences (≥10 identical reads) of control samples of A 2 (A; blue), subtypes D 1 (B; green), a mixture of A 2 and D 1 at a 50:50 ratio (C; red), and a merged tree of all three sample runs (D) are shown. The trees were constructed with a selection of subtype reference sequences and random sequences from individuals in Rakai shown in black. Brackets demonstrate the source of different clades within the merged trees. Bootstrap values higher than 80% are shown for nonmerged trees (1,000 replicates).

    Techniques Used: Next-Generation Sequencing, Construct, Selection

    10) Product Images from "Comparable response of ccn1 with ccn2 genes upon arthritis: An in vitro evaluation with a human chondrocytic cell line stimulated by a set of cytokines"

    Article Title: Comparable response of ccn1 with ccn2 genes upon arthritis: An in vitro evaluation with a human chondrocytic cell line stimulated by a set of cytokines

    Journal: Cell communication and signaling : CCS

    doi: 10.1186/1478-811X-3-6

    Distinctive quantification of ccn1 and ccn2 mRNA by real-time RT-PCR . A. Primers used for real-time PCR and the structures of human ccn1 and ccn2 mRNAs. Schematic representations are illustrated in reference to the modular structure of human CCN1 and CCN2 (stippled boxes). The small open circle and AAAAA at the left and right ends denote the 5'-cap structure and poly-A tail, respectively. Names, locations for recognition, nucleotide sequence of the primers, and the expected sizes of the PCR products are given. Abbreviations: IGFBP, insulin-like growth factor binding module; VWC, von Willebrand factor type C module; TSP1, thrombospondin type 1 repeat; CT, C-terminal module. B. The CCN1 (219 bp), CCN2 (153 bp), and β-actin (101 bp) PCR products amplified by LightCycler were analyzed by agarose electrophoresis. C. Melting curve analysis of the RT- PCR products of ccn1 and ccn2 . Melting curves were acquired after 45 cycles of amplification. Melting curve pattern is displayed on the left panel, where fluorescent intensity (F1) from SYBR green is plotted against temperature. Melting peak pattern can be found on the right panel, in which the decrement of F1 is plotted against temperature.
    Figure Legend Snippet: Distinctive quantification of ccn1 and ccn2 mRNA by real-time RT-PCR . A. Primers used for real-time PCR and the structures of human ccn1 and ccn2 mRNAs. Schematic representations are illustrated in reference to the modular structure of human CCN1 and CCN2 (stippled boxes). The small open circle and AAAAA at the left and right ends denote the 5'-cap structure and poly-A tail, respectively. Names, locations for recognition, nucleotide sequence of the primers, and the expected sizes of the PCR products are given. Abbreviations: IGFBP, insulin-like growth factor binding module; VWC, von Willebrand factor type C module; TSP1, thrombospondin type 1 repeat; CT, C-terminal module. B. The CCN1 (219 bp), CCN2 (153 bp), and β-actin (101 bp) PCR products amplified by LightCycler were analyzed by agarose electrophoresis. C. Melting curve analysis of the RT- PCR products of ccn1 and ccn2 . Melting curves were acquired after 45 cycles of amplification. Melting curve pattern is displayed on the left panel, where fluorescent intensity (F1) from SYBR green is plotted against temperature. Melting peak pattern can be found on the right panel, in which the decrement of F1 is plotted against temperature.

    Techniques Used: Quantitative RT-PCR, Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction, Binding Assay, Amplification, Electrophoresis, Reverse Transcription Polymerase Chain Reaction, SYBR Green Assay

    11) Product Images from "Endothelial LRP1 regulates metabolic responses by acting as a co-activator of PPARγ"

    Article Title: Endothelial LRP1 regulates metabolic responses by acting as a co-activator of PPARγ

    Journal: Nature Communications

    doi: 10.1038/ncomms14960

    Lrp1β binds to Pparγ and promotes its transcriptional activity. ( a ) Lysates of HEK 293 cells with overexpressed Flag-tagged Pparγ (Flag-Pparγ) and HA-tagged mini-Lrp1 receptor (HA-mLrp1 13 ) were immunoprecipitated (IPed) and blotted with the indicated antibodies. ( b ) GST pull-down assay was performed with recombinant Pparγ protein. GST-Lrp1-ICD: GST-tagged Lrp1 ICD (GST-ICD; a.a. 4445–4544 of human Lrp1). ( c ) Co-IP assay was performed with lysates of HEK293 cells that were transfected with the indicated constructs. ( d ) Ppar reporter assay was performed with lysates of HEK293 cells that were treated with 10 μM pioglitazone (Piog). ( e , f ) In vivo PPRE-driven luciferase activity was evaluated with Lrp1 f/f ;CAG-CreER +/− ;PPRE-luc + (CAG-Cre+;PPRE-luc+ and CAG-Cre−;PPRE-luc+ generated by the breeding of B6;129S7- Lrp1 tm2Her /J, B6.Cg-Tg(CAG-cre/Esr1*)5Amc/J and repTOP PPRE-Luc mice) mice. Pioglitazone, 50 or 150 mg kg −1 . Images presented in e are representative results of the indicated mouse group and bioluminescence intensity (BLI) was quantified in f . ( g ) Pdk4 mRNA level was measured with real-time PCR assays. Lrp1 knockout MEFs (Lrp1 −/− MEFs) and Wt (Lrp1 +/+ MEFs) were treated with 10 μM pioglitazone. ( h ) ChIP-qPCR was performed for endogenous Pdk4 promoter and negative DNA region control (Neg). Stable Lrp1β-transfected HEK293 cells were treated with pioglitazone (Piog) at 10 μM or palmitic acids (PA) at 0.5 mM for 24 h. Enriched chromatin fractions were IPed with Lrp1 or IgG. ( i , j ) HEK293 cells were transfected with Flag-Lrp1β-Wt (VGGLL) or Mut (VGGAA). Their lysates were mixed with GST fused recombinant Pparγ LBD (Pparγ-LBD) protein for IP and immunoblotting. The bound Pparγ-LBD protein level was quantified by measuring its band intensity and normalized by the amount of enriched Flag-Lrp1β protein ( j ). ( k ) Ppar reporter assay was performed in HEK293 cells. ( l ) HEK293 cells were transfected with Flag-tagged Lrp1β-Wt or Mut and ChIP assays were performed with Flag antibody or IgG. n =3 for cell culture experiments in a – d , g – l . n =3 for Lrp1 f/f ;CAG-CreER + ;PPRE-luc + mice and 4 for Lrp1 f/f ;CAG-CreER − ;PPRE-luc + mice in e , f . * P
    Figure Legend Snippet: Lrp1β binds to Pparγ and promotes its transcriptional activity. ( a ) Lysates of HEK 293 cells with overexpressed Flag-tagged Pparγ (Flag-Pparγ) and HA-tagged mini-Lrp1 receptor (HA-mLrp1 13 ) were immunoprecipitated (IPed) and blotted with the indicated antibodies. ( b ) GST pull-down assay was performed with recombinant Pparγ protein. GST-Lrp1-ICD: GST-tagged Lrp1 ICD (GST-ICD; a.a. 4445–4544 of human Lrp1). ( c ) Co-IP assay was performed with lysates of HEK293 cells that were transfected with the indicated constructs. ( d ) Ppar reporter assay was performed with lysates of HEK293 cells that were treated with 10 μM pioglitazone (Piog). ( e , f ) In vivo PPRE-driven luciferase activity was evaluated with Lrp1 f/f ;CAG-CreER +/− ;PPRE-luc + (CAG-Cre+;PPRE-luc+ and CAG-Cre−;PPRE-luc+ generated by the breeding of B6;129S7- Lrp1 tm2Her /J, B6.Cg-Tg(CAG-cre/Esr1*)5Amc/J and repTOP PPRE-Luc mice) mice. Pioglitazone, 50 or 150 mg kg −1 . Images presented in e are representative results of the indicated mouse group and bioluminescence intensity (BLI) was quantified in f . ( g ) Pdk4 mRNA level was measured with real-time PCR assays. Lrp1 knockout MEFs (Lrp1 −/− MEFs) and Wt (Lrp1 +/+ MEFs) were treated with 10 μM pioglitazone. ( h ) ChIP-qPCR was performed for endogenous Pdk4 promoter and negative DNA region control (Neg). Stable Lrp1β-transfected HEK293 cells were treated with pioglitazone (Piog) at 10 μM or palmitic acids (PA) at 0.5 mM for 24 h. Enriched chromatin fractions were IPed with Lrp1 or IgG. ( i , j ) HEK293 cells were transfected with Flag-Lrp1β-Wt (VGGLL) or Mut (VGGAA). Their lysates were mixed with GST fused recombinant Pparγ LBD (Pparγ-LBD) protein for IP and immunoblotting. The bound Pparγ-LBD protein level was quantified by measuring its band intensity and normalized by the amount of enriched Flag-Lrp1β protein ( j ). ( k ) Ppar reporter assay was performed in HEK293 cells. ( l ) HEK293 cells were transfected with Flag-tagged Lrp1β-Wt or Mut and ChIP assays were performed with Flag antibody or IgG. n =3 for cell culture experiments in a – d , g – l . n =3 for Lrp1 f/f ;CAG-CreER + ;PPRE-luc + mice and 4 for Lrp1 f/f ;CAG-CreER − ;PPRE-luc + mice in e , f . * P

    Techniques Used: Activity Assay, Immunoprecipitation, Pull Down Assay, Recombinant, Co-Immunoprecipitation Assay, Transfection, Construct, Reporter Assay, In Vivo, Luciferase, Generated, Mouse Assay, Real-time Polymerase Chain Reaction, Knock-Out, Chromatin Immunoprecipitation, Cell Culture

    12) Product Images from "Accuracy, Precision, and Method Detection Limits of Quantitative PCR for Airborne Bacteria and Fungi ▿"

    Article Title: Accuracy, Precision, and Method Detection Limits of Quantitative PCR for Airborne Bacteria and Fungi ▿

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.01240-10

    Instrument repeatability as a function of cell quantity COV ( n = 7). The qPCR MDLs are 5 cells for B. atrophaeus and 0.05 cells for A. fumigatus .
    Figure Legend Snippet: Instrument repeatability as a function of cell quantity COV ( n = 7). The qPCR MDLs are 5 cells for B. atrophaeus and 0.05 cells for A. fumigatus .

    Techniques Used: Real-time Polymerase Chain Reaction

    13) Product Images from "Genome-Wide Prediction and Validation of Intergenic Enhancers in Arabidopsis Using Open Chromatin Signatures [OPEN]"

    Article Title: Genome-Wide Prediction and Validation of Intergenic Enhancers in Arabidopsis Using Open Chromatin Signatures [OPEN]

    Journal: The Plant Cell

    doi: 10.1105/tpc.15.00537

    DNase I-PCR Assay of the DNase I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .
    Figure Legend Snippet: DNase I-PCR Assay of the DNase I Sensitivity of a Transgenic Enhancer Sequence. (A) . (B) A diagram illustrating the design of a PCR primer that allows the transgenic L3 locus, but not the endogenous L3, to be specifically amplified. (C) DNase I-PCR shows differential DNase I sensitivity of chromatin in selected genomic regions. The DNase I sensitivity of the transgenic L3 locus was similar to that of the open chromatin control ACTIN7 .

    Techniques Used: Polymerase Chain Reaction, Transgenic Assay, Sequencing, Amplification

    14) Product Images from "An Upstream Open Reading Frame and the Context of the Two AUG Codons Affect the Abundance of Mitochondrial and Nuclear RNase H1 ▿An Upstream Open Reading Frame and the Context of the Two AUG Codons Affect the Abundance of Mitochondrial and Nuclear RNase H1 ▿ †"

    Article Title: An Upstream Open Reading Frame and the Context of the Two AUG Codons Affect the Abundance of Mitochondrial and Nuclear RNase H1 ▿An Upstream Open Reading Frame and the Context of the Two AUG Codons Affect the Abundance of Mitochondrial and Nuclear RNase H1 ▿ †

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.00619-10

    Overexpression of RNase H1 in mitochondria impairs growth and results in mtDNA loss. (A) FT293-CAT, FT293-M27, FT293-M1-M27I, and FT293-M1 cells were grown in the media with (filled squares) or without (open squares) 1 μg/ml doxycycline. The FT293-CAT cell line has the CAT gene inserted at the flip-in site. Likewise, the other FT293 cell lines have the cDNA for the Rnaseh1 mRNA, starting at M1 (FT293-M1), M27 (FT293-M27), or M1 with the M27 AUG changed to AUC (FT293-M1-M27I). The numbers of cells were counted with a hemocytometer. (B) RNase H1-HA-NESi is expressed, activated, and localized to mitochondria in 293 cells. RNase H1 mRNA levels were assayed by RT-qPCR at different time points and dosages of doxycycline. The initial black bars are the parental FT293 cell line lacking RNase H1-HA-NESi. Subsequent bars from left to right in each set are cells with RNase H-HA-NESi expressed at 0 (red), 1 (blue), 3 (yellow), and 10 (green) ng/ml doxycycline. Protein contents of cell lysates were analyzed by Coomassie brilliant blue (CBB) stain, Western blotting for the HA epitope, and the RNase H1 gel activity assay with poly(rA)/poly(dT) as the substrate. Equal amounts of total protein (as assessed from the CBB staining) were loaded in each lane. For clarity, activity is shown as a negative image. Expression of RNase H1-HA-NESi was induced for 6 days using 0, 3, 30, and 60 ng/ml doxycycline. At the bottom and to the right is shown confocal microscopy of FT293-RNase H1-HA-NESi. After 24 h of induction using 100 ng/ml doxycycline, the entire signal detected with the anti-HA antibody (green) colocalized with mitochondria (MitoTracker orange). mtDNA copy number decreases as the dosage of doxycycline and days of incubation increase. For each day, the first bar (black) is FT293 without the cDNA for human RNase H1 treated with 10 ng/ml doxycycline. Subsequent bars from left to right in each set contain 0 (violet), 0.1 (red), 1 (blue), 3 (yellow), or 10 (green) ng/ml doxycycline. At day 6, the values for 1, 3, and 10 ng/ml doxycycline are 0.036, 0.032, and 0.027, respectively. At day 9, the values for 1, 3, and 10 ng/ml doxycycline are 0.018, 0.011, and 0.010, respectively.
    Figure Legend Snippet: Overexpression of RNase H1 in mitochondria impairs growth and results in mtDNA loss. (A) FT293-CAT, FT293-M27, FT293-M1-M27I, and FT293-M1 cells were grown in the media with (filled squares) or without (open squares) 1 μg/ml doxycycline. The FT293-CAT cell line has the CAT gene inserted at the flip-in site. Likewise, the other FT293 cell lines have the cDNA for the Rnaseh1 mRNA, starting at M1 (FT293-M1), M27 (FT293-M27), or M1 with the M27 AUG changed to AUC (FT293-M1-M27I). The numbers of cells were counted with a hemocytometer. (B) RNase H1-HA-NESi is expressed, activated, and localized to mitochondria in 293 cells. RNase H1 mRNA levels were assayed by RT-qPCR at different time points and dosages of doxycycline. The initial black bars are the parental FT293 cell line lacking RNase H1-HA-NESi. Subsequent bars from left to right in each set are cells with RNase H-HA-NESi expressed at 0 (red), 1 (blue), 3 (yellow), and 10 (green) ng/ml doxycycline. Protein contents of cell lysates were analyzed by Coomassie brilliant blue (CBB) stain, Western blotting for the HA epitope, and the RNase H1 gel activity assay with poly(rA)/poly(dT) as the substrate. Equal amounts of total protein (as assessed from the CBB staining) were loaded in each lane. For clarity, activity is shown as a negative image. Expression of RNase H1-HA-NESi was induced for 6 days using 0, 3, 30, and 60 ng/ml doxycycline. At the bottom and to the right is shown confocal microscopy of FT293-RNase H1-HA-NESi. After 24 h of induction using 100 ng/ml doxycycline, the entire signal detected with the anti-HA antibody (green) colocalized with mitochondria (MitoTracker orange). mtDNA copy number decreases as the dosage of doxycycline and days of incubation increase. For each day, the first bar (black) is FT293 without the cDNA for human RNase H1 treated with 10 ng/ml doxycycline. Subsequent bars from left to right in each set contain 0 (violet), 0.1 (red), 1 (blue), 3 (yellow), or 10 (green) ng/ml doxycycline. At day 6, the values for 1, 3, and 10 ng/ml doxycycline are 0.036, 0.032, and 0.027, respectively. At day 9, the values for 1, 3, and 10 ng/ml doxycycline are 0.018, 0.011, and 0.010, respectively.

    Techniques Used: Over Expression, Quantitative RT-PCR, Staining, Western Blot, Activity Assay, Expressing, Confocal Microscopy, Incubation

    15) Product Images from "Local overexpression of the myostatin propeptide increases glucose transporter expression and enhances skeletal muscle glucose disposal"

    Article Title: Local overexpression of the myostatin propeptide increases glucose transporter expression and enhances skeletal muscle glucose disposal

    Journal: American Journal of Physiology - Endocrinology and Metabolism

    doi: 10.1152/ajpendo.00586.2013

    Effect of myostatin blockade on selected mediators of muscle size and metabolism. Total RNA and cDNA was prepared from paired TC muscles, and SYBR Green Real-time PCR was conducted using 20 ng of each cDNA or DNA standard in duplicate and primers as listed in materials and methods . Summary data (means ± SE; n = 10–12) are shown for each target mRNA of interest [insulin-like growth factor-I (IGF-I), IGF-I receptor (IGF-I Rec), FOXO3, peroxisome proliferator-activated receptor coactivator 1α (PGC1a), nuclear factor-κB p65 subunit (p65), activin IIB receptor (activin 2BR), latent transforming growth factor-β-binding protein (LTBP3), muscle ring finger protein 1 (MURF1), and atrogin 1], normalized to the geometric mean of the reference mRNAs cyclophilin, 36B4, and 18S. Filled bars, control; open bars, test muscle. * P
    Figure Legend Snippet: Effect of myostatin blockade on selected mediators of muscle size and metabolism. Total RNA and cDNA was prepared from paired TC muscles, and SYBR Green Real-time PCR was conducted using 20 ng of each cDNA or DNA standard in duplicate and primers as listed in materials and methods . Summary data (means ± SE; n = 10–12) are shown for each target mRNA of interest [insulin-like growth factor-I (IGF-I), IGF-I receptor (IGF-I Rec), FOXO3, peroxisome proliferator-activated receptor coactivator 1α (PGC1a), nuclear factor-κB p65 subunit (p65), activin IIB receptor (activin 2BR), latent transforming growth factor-β-binding protein (LTBP3), muscle ring finger protein 1 (MURF1), and atrogin 1], normalized to the geometric mean of the reference mRNAs cyclophilin, 36B4, and 18S. Filled bars, control; open bars, test muscle. * P

    Techniques Used: SYBR Green Assay, Real-time Polymerase Chain Reaction, Binding Assay

    16) Product Images from "IL-12p40 Homodimer Ameliorates Experimental Autoimmune Arthritis"

    Article Title: IL-12p40 Homodimer Ameliorates Experimental Autoimmune Arthritis

    Journal: The Journal of Immunology Author Choice

    doi: 10.4049/jimmunol.1500400

    (p40) 2 inhibits inflammatory cytokine expression in IL-1RaKO mice. ( A – G ) All tissue and cells were obtained from therapeutically treated IL-1RaKO mice. (A) IL-1RaKO mice joint tissue was stained with mAb species for IL-23, IL-12p70, IL-17, IFN-γ, IL-1β, TNF-α, and IL-6 (obtained at 6 wk). Brown represents positive staining for IL-23, IL-12p70, Il-17, IFN-γ, IL-1β, TNF-α, and IL-6 (original magnification ×200). Data shown are representative of three independent experiments. (B) Spleen cells of wild-type (WT), mock vector, (p40) 2 vector, and PBS mice were harvested at the peak of disease (obtained at 6 wk). mRNA expression of IL-23p19 and IL-12 was analyzed by real-time PCR. (C and D) IL-23p19 and IL-12 production was analyzed using ELISA from the culture supernatants of each group. (E) mRNA expression of IL-1β, TNF-α, IL-6, IL-17, and IFN-γ was analyzed by real-time PCR in joint cells. (F and G) Joint cells of the (p40) 2 injection group and control group were cultured with IL-23 and IL-12, with or without (p40) 2 , for 3 d. mRNA expression of IL-17, IFN-γ, IL-1β, and TNF-α was assessed by real-time PCR. Data are mean ± SD and are representative of three independent experiments. * p
    Figure Legend Snippet: (p40) 2 inhibits inflammatory cytokine expression in IL-1RaKO mice. ( A – G ) All tissue and cells were obtained from therapeutically treated IL-1RaKO mice. (A) IL-1RaKO mice joint tissue was stained with mAb species for IL-23, IL-12p70, IL-17, IFN-γ, IL-1β, TNF-α, and IL-6 (obtained at 6 wk). Brown represents positive staining for IL-23, IL-12p70, Il-17, IFN-γ, IL-1β, TNF-α, and IL-6 (original magnification ×200). Data shown are representative of three independent experiments. (B) Spleen cells of wild-type (WT), mock vector, (p40) 2 vector, and PBS mice were harvested at the peak of disease (obtained at 6 wk). mRNA expression of IL-23p19 and IL-12 was analyzed by real-time PCR. (C and D) IL-23p19 and IL-12 production was analyzed using ELISA from the culture supernatants of each group. (E) mRNA expression of IL-1β, TNF-α, IL-6, IL-17, and IFN-γ was analyzed by real-time PCR in joint cells. (F and G) Joint cells of the (p40) 2 injection group and control group were cultured with IL-23 and IL-12, with or without (p40) 2 , for 3 d. mRNA expression of IL-17, IFN-γ, IL-1β, and TNF-α was assessed by real-time PCR. Data are mean ± SD and are representative of three independent experiments. * p

    Techniques Used: Expressing, Mouse Assay, Staining, Plasmid Preparation, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Injection, Cell Culture

    (p40) 2 inhibits Ag-specific T cell proliferation and cytokine production in CIA. ( A – C ) All cells were obtained from (p40) 2 -treated or mock-treated CIA mice (obtained at 5 wk). (A) CII-reactive T cells (1 × 10 5 /ml) and irradiated (2500 rad) APCs (1 × 10 5 /ml) isolated from spleens of CIA mice, (p40) 2 -treated mice, or mock-treated mice. The cells were cocultured for 3 d. (B) T cells and irradiated APCs isolated from CIA mice, (p40) 2 -treated mice, or mock-treated mice were cocultured with CII, OVA, and CII plus (p40) 2 for 3 d, and CD4 + T cell proliferation was measured. Data are mean cpm of triplicate cultures. (C) The culture supernatant was measured by ELISA for IL-23, IL-17, IL-1β, and TNF-α. Data are mean ± SD and are representative of three independent experiments. * p and # p
    Figure Legend Snippet: (p40) 2 inhibits Ag-specific T cell proliferation and cytokine production in CIA. ( A – C ) All cells were obtained from (p40) 2 -treated or mock-treated CIA mice (obtained at 5 wk). (A) CII-reactive T cells (1 × 10 5 /ml) and irradiated (2500 rad) APCs (1 × 10 5 /ml) isolated from spleens of CIA mice, (p40) 2 -treated mice, or mock-treated mice. The cells were cocultured for 3 d. (B) T cells and irradiated APCs isolated from CIA mice, (p40) 2 -treated mice, or mock-treated mice were cocultured with CII, OVA, and CII plus (p40) 2 for 3 d, and CD4 + T cell proliferation was measured. Data are mean cpm of triplicate cultures. (C) The culture supernatant was measured by ELISA for IL-23, IL-17, IL-1β, and TNF-α. Data are mean ± SD and are representative of three independent experiments. * p and # p

    Techniques Used: Mouse Assay, Irradiation, Isolation, Enzyme-linked Immunosorbent Assay

    17) Product Images from "Cold tolerance and silencing of three cold-tolerance genes of overwintering Chinese white pine larvae"

    Article Title: Cold tolerance and silencing of three cold-tolerance genes of overwintering Chinese white pine larvae

    Journal: Scientific Reports

    doi: 10.1038/srep34698

    qRT-PCR analysis of DarmSDH, DarmTPS and DarmGLK transcript patterns from D. armandi larvae; after injected for 24 h, 48 h and 72 h. The standard errors of the means of three biological replicates are represented by error bars. Transcript patterns of DarmSDH and DarmTPS were analyzed on December 2014, and DarmGLK on November 2014.
    Figure Legend Snippet: qRT-PCR analysis of DarmSDH, DarmTPS and DarmGLK transcript patterns from D. armandi larvae; after injected for 24 h, 48 h and 72 h. The standard errors of the means of three biological replicates are represented by error bars. Transcript patterns of DarmSDH and DarmTPS were analyzed on December 2014, and DarmGLK on November 2014.

    Techniques Used: Quantitative RT-PCR, Injection

    18) Product Images from "The influence of CYP3A, PPARA, and POR genetic variants on the pharmacokinetics of tacrolimus and cyclosporine in renal transplant recipients"

    Article Title: The influence of CYP3A, PPARA, and POR genetic variants on the pharmacokinetics of tacrolimus and cyclosporine in renal transplant recipients

    Journal: European Journal of Clinical Pharmacology

    doi: 10.1007/s00228-014-1656-3

    Cyclosporine C 2 /D ratio (μg*L -1 /mg) as a function of CYP3A5*3, CYP3A4*22, PPARA c.209-1003G > A , PPARA c.208 + 3819A > G, and POR*28 . The box-and-whisker plots indicate interquartile ranges ( boxes ), medians ( horizontal lines in the boxes ), and highest and lowest values ( whiskers above and below the boxes ). P values are related to the ANOVA test, as described under Results . C 2 /D, dose-adjusted concentrations before dosing; CYP, gene encoding cytochrome P450; PPARA, gene encoding the nuclear receptor peroxisome proliferator-activated receptor alpha; POR, gene encoding cytochrome P450 oxidoreductase
    Figure Legend Snippet: Cyclosporine C 2 /D ratio (μg*L -1 /mg) as a function of CYP3A5*3, CYP3A4*22, PPARA c.209-1003G > A , PPARA c.208 + 3819A > G, and POR*28 . The box-and-whisker plots indicate interquartile ranges ( boxes ), medians ( horizontal lines in the boxes ), and highest and lowest values ( whiskers above and below the boxes ). P values are related to the ANOVA test, as described under Results . C 2 /D, dose-adjusted concentrations before dosing; CYP, gene encoding cytochrome P450; PPARA, gene encoding the nuclear receptor peroxisome proliferator-activated receptor alpha; POR, gene encoding cytochrome P450 oxidoreductase

    Techniques Used: Whisker Assay

    Tacrolimus C 0 /D ratio (μg*L -1 /mg) as a function of CYP3A5*3, CYP3A4*22, PPARA c.209-1003G > A , PPARA c.208 + 3819A > G , and POR*28 . The box-and-whisker plots indicate interquartile ranges ( boxes ), medians ( horizontal lines in the boxes ), and the highest and lowest values ( whiskers above and below the boxes ). P values are related to the ANOVA test, described under Results . C 0 /D, dose-adjusted concentrations before dosing; CYP, gene encoding cytochrome P450; PPARA, gene encoding the nuclear receptor peroxisome proliferator-activated receptor alpha; POR, gene encoding cytochrome P450 oxidoreductase
    Figure Legend Snippet: Tacrolimus C 0 /D ratio (μg*L -1 /mg) as a function of CYP3A5*3, CYP3A4*22, PPARA c.209-1003G > A , PPARA c.208 + 3819A > G , and POR*28 . The box-and-whisker plots indicate interquartile ranges ( boxes ), medians ( horizontal lines in the boxes ), and the highest and lowest values ( whiskers above and below the boxes ). P values are related to the ANOVA test, described under Results . C 0 /D, dose-adjusted concentrations before dosing; CYP, gene encoding cytochrome P450; PPARA, gene encoding the nuclear receptor peroxisome proliferator-activated receptor alpha; POR, gene encoding cytochrome P450 oxidoreductase

    Techniques Used: Whisker Assay

    19) Product Images from "Uterine extracellular matrix components are altered during defective decidualization in interleukin-11 receptor ? deficient mice"

    Article Title: Uterine extracellular matrix components are altered during defective decidualization in interleukin-11 receptor ? deficient mice

    Journal: Reproductive biology and endocrinology : RB & E

    doi: 10.1186/1477-7827-2-76

    Gene expression in IL11Ra +/+ and IL11Ra -/- uterus following artificial decidualization. Expression profiling of 15K genes between IL11Ra +/+ and IL11Ra -/- at 0 h (A, B), 18 h (C, D), 24 h (E, F) and 48 h (G, H) following the artificial induction of decidualization. Each volcano style plot shows the normalized log ratio (effect estimate) of IL11Ra -/- compared to wild type for each gene from a series of 4 microarrays, plotted against the log odds of differential expression. A, C, E, G represent the first replicates, and B, D, F, H the second dye-swapped replicates. Genes with log odds of differential expression greater than 3 (ie. adjusted p -value
    Figure Legend Snippet: Gene expression in IL11Ra +/+ and IL11Ra -/- uterus following artificial decidualization. Expression profiling of 15K genes between IL11Ra +/+ and IL11Ra -/- at 0 h (A, B), 18 h (C, D), 24 h (E, F) and 48 h (G, H) following the artificial induction of decidualization. Each volcano style plot shows the normalized log ratio (effect estimate) of IL11Ra -/- compared to wild type for each gene from a series of 4 microarrays, plotted against the log odds of differential expression. A, C, E, G represent the first replicates, and B, D, F, H the second dye-swapped replicates. Genes with log odds of differential expression greater than 3 (ie. adjusted p -value

    Techniques Used: Expressing

    Uterine weight following artificial decidualization. Weight (mg +/- SEM) of uterine horns at times following artificial decidualization of IL11Ra +/+ (open bars) and IL11Ra -/- (shaded bars) littermates. ** p
    Figure Legend Snippet: Uterine weight following artificial decidualization. Weight (mg +/- SEM) of uterine horns at times following artificial decidualization of IL11Ra +/+ (open bars) and IL11Ra -/- (shaded bars) littermates. ** p

    Techniques Used:

    Immunohistochemistry for extracellular matrix components. Immunohistochemical staining of wild type (A, C, E, G, I, K, M, O, P, Q, S) and IL11Ra -/- (B, D, F, H, J, L, N, R, T) uterus at 48 h of decidualization using specific antibodies for collagen III (A, B, C, D), biglycan (E, F, G, H), nidogen-1 (I, J, K, L), SPARC (M, N, O, P) and desmin (Q, R, S, T). Negative controls using a matching concentration of non-immune IgG (collagen III, nidogen-1, SPARC and desmin) or normal serum (biglycan) in place of the primary antibody are inset in A, B, E, F, I, J, M, N, Q and R. Black squares on A and B indicate the antimesometrial pole magnified in C and D. Abbreviations: connective tissue (ct), myometrium (my), mesometrial pole (m), antimesometrial pole (am), luminal epithelium (le), glandular epithelium (ge), decidualized stromal cell (dsc), non-decidualized stromal cell (sc), blood vessel (bv), glycocalyx (gly). Scale bar = 50 μm (A, B, E, F, I, J, M, N, Q and R are at the same magnification; C, D, G and P are at the same magnification; H, K, L, O, S, T and inset in G are at the same magnification).
    Figure Legend Snippet: Immunohistochemistry for extracellular matrix components. Immunohistochemical staining of wild type (A, C, E, G, I, K, M, O, P, Q, S) and IL11Ra -/- (B, D, F, H, J, L, N, R, T) uterus at 48 h of decidualization using specific antibodies for collagen III (A, B, C, D), biglycan (E, F, G, H), nidogen-1 (I, J, K, L), SPARC (M, N, O, P) and desmin (Q, R, S, T). Negative controls using a matching concentration of non-immune IgG (collagen III, nidogen-1, SPARC and desmin) or normal serum (biglycan) in place of the primary antibody are inset in A, B, E, F, I, J, M, N, Q and R. Black squares on A and B indicate the antimesometrial pole magnified in C and D. Abbreviations: connective tissue (ct), myometrium (my), mesometrial pole (m), antimesometrial pole (am), luminal epithelium (le), glandular epithelium (ge), decidualized stromal cell (dsc), non-decidualized stromal cell (sc), blood vessel (bv), glycocalyx (gly). Scale bar = 50 μm (A, B, E, F, I, J, M, N, Q and R are at the same magnification; C, D, G and P are at the same magnification; H, K, L, O, S, T and inset in G are at the same magnification).

    Techniques Used: Immunohistochemistry, Staining, Concentration Assay

    20) Product Images from "hsp70-Dependent Antiviral Immunity against Cytopathic Neuronal Infection by Vesicular Stomatitis Virus"

    Article Title: hsp70-Dependent Antiviral Immunity against Cytopathic Neuronal Infection by Vesicular Stomatitis Virus

    Journal: Journal of Virology

    doi: 10.1128/JVI.00872-13

    Protection mediated by hsp70 that is expressed from the VSV genome is correlated with enhanced induction of IFN-β transcripts in brain. NT mice ( n = 4 per group) received intranasal inoculations of 1 × 10 6 PFU rVSV-hsp70, rVSV-VP1, or rVSV (parent virus). Mice injected with PBS served as uninfected controls. Total brain RNA was extracted from the rostral brain at 1 day p.i., and IFN-β transcript levels were determined using real-time RT-qPCR. Results were directly compared to levels of IFN-β measured in brains of TG mice infected with rVSV and in uninfected TG controls. Significantly increased levels of IFN-β transcript were observed in rVSV-hsp70-infected NT mice relative to rVSV-VP1-infected (Inf), rVSV-infected, and uninfected (Uninf) NT mice ( P
    Figure Legend Snippet: Protection mediated by hsp70 that is expressed from the VSV genome is correlated with enhanced induction of IFN-β transcripts in brain. NT mice ( n = 4 per group) received intranasal inoculations of 1 × 10 6 PFU rVSV-hsp70, rVSV-VP1, or rVSV (parent virus). Mice injected with PBS served as uninfected controls. Total brain RNA was extracted from the rostral brain at 1 day p.i., and IFN-β transcript levels were determined using real-time RT-qPCR. Results were directly compared to levels of IFN-β measured in brains of TG mice infected with rVSV and in uninfected TG controls. Significantly increased levels of IFN-β transcript were observed in rVSV-hsp70-infected NT mice relative to rVSV-VP1-infected (Inf), rVSV-infected, and uninfected (Uninf) NT mice ( P

    Techniques Used: Mouse Assay, Injection, Quantitative RT-PCR, Infection

    21) Product Images from "Adjusting microbiome profiles for differences in microbial load by spike-in bacteria"

    Article Title: Adjusting microbiome profiles for differences in microbial load by spike-in bacteria

    Journal: Microbiome

    doi: 10.1186/s40168-016-0175-0

    Procedural overview of proposed spike-in procedure and the spike-in-based calibration to total microbial load (SCML). The overview is divided into four sections: spike-in procedure and bacterial lysis (blue), DNA isolation, amplification and sequencing (yellow), pre-processing (red) and the actual spike-in-based calibration to microbial load (green). White-filled boxes depict procedural intermediates, while grey-filled boxes depict the different procedural steps. Each step is numbered. In the first step (1) whole cells of exogenous spike bacteria corresponding to a fixed number of 16S rDNA copies are added to homogenized microbiome samples. Bacterial lysis is performed on the resulting spiked samples (2). Metagenomic DNA is extracted from the lysates (3) and PCR amplified using 16S rDNA specific primers (4), creating 16S rDNA amplicons. These amplicons are purified and pyrosequencing is performed (5). The resulting raw read counts are pre-processed with QIIME (quality filtering, demultiplexing and closed reference OTU picking) to generate OTU read count tables (6). Based on the read counts associated with single or multiple reference spike-in bacteria, a size factor s i for each sample i is calculated and applied to each OTU of this particular sample i (8, see methods section). This leads to an OTU read count table calibrated to differences in microbial load. These read counts can be utilized to more accurately assess changes between different samples. All depicted steps are described in detail in the methods section. Stars indicate points in the procedure at which qPCR is performed to identify possible errors in DNA isolation (metagenomic DNA) or PCR amplification (16S rDNA amplicons).
    Figure Legend Snippet: Procedural overview of proposed spike-in procedure and the spike-in-based calibration to total microbial load (SCML). The overview is divided into four sections: spike-in procedure and bacterial lysis (blue), DNA isolation, amplification and sequencing (yellow), pre-processing (red) and the actual spike-in-based calibration to microbial load (green). White-filled boxes depict procedural intermediates, while grey-filled boxes depict the different procedural steps. Each step is numbered. In the first step (1) whole cells of exogenous spike bacteria corresponding to a fixed number of 16S rDNA copies are added to homogenized microbiome samples. Bacterial lysis is performed on the resulting spiked samples (2). Metagenomic DNA is extracted from the lysates (3) and PCR amplified using 16S rDNA specific primers (4), creating 16S rDNA amplicons. These amplicons are purified and pyrosequencing is performed (5). The resulting raw read counts are pre-processed with QIIME (quality filtering, demultiplexing and closed reference OTU picking) to generate OTU read count tables (6). Based on the read counts associated with single or multiple reference spike-in bacteria, a size factor s i for each sample i is calculated and applied to each OTU of this particular sample i (8, see methods section). This leads to an OTU read count table calibrated to differences in microbial load. These read counts can be utilized to more accurately assess changes between different samples. All depicted steps are described in detail in the methods section. Stars indicate points in the procedure at which qPCR is performed to identify possible errors in DNA isolation (metagenomic DNA) or PCR amplification (16S rDNA amplicons).

    Techniques Used: Lysis, DNA Extraction, Amplification, Sequencing, Polymerase Chain Reaction, Purification, Real-time Polymerase Chain Reaction

    Comparison of SCML and normalization by qRT-PCR-derived total number of 16S rDNA copies to all background OTUs. Observed log 2 ratio versus expected log 2 ratio of all background bacteria OTUs for all pairwise sample comparisons after ( a ) SCML by S. ruber and ( b ) normalization by qRT-PCR derived total 16S rDNA copy number. The data is binned to hexagons because of the high number of data points. The colour of each hexagon represents the percentage of all counts at the corresponding level of expected log 2 ratios contained in each bin. Bins that contributed to less than 0.05 percent for each level of expected log 2 ratio are omitted. The purple diagonal represents the identity, which represents the expected log 2 ratios by design. The box-plots in ( c ) summarize the error between the expected and observed log 2 ratios for the four different approaches. The smaller this error, the better calibrated the ratios are. Variances of the log 2 differences are 3.65, 2.01, 1.28 and 1.18 as derived from relative abundances, counts calibrated for differences in total number of 16S rRNA gene copies, SCML (by S. ruber ) and combined SCML (by S. ruber , A. acidiphilus and R. radiobacter ), respectively
    Figure Legend Snippet: Comparison of SCML and normalization by qRT-PCR-derived total number of 16S rDNA copies to all background OTUs. Observed log 2 ratio versus expected log 2 ratio of all background bacteria OTUs for all pairwise sample comparisons after ( a ) SCML by S. ruber and ( b ) normalization by qRT-PCR derived total 16S rDNA copy number. The data is binned to hexagons because of the high number of data points. The colour of each hexagon represents the percentage of all counts at the corresponding level of expected log 2 ratios contained in each bin. Bins that contributed to less than 0.05 percent for each level of expected log 2 ratio are omitted. The purple diagonal represents the identity, which represents the expected log 2 ratios by design. The box-plots in ( c ) summarize the error between the expected and observed log 2 ratios for the four different approaches. The smaller this error, the better calibrated the ratios are. Variances of the log 2 differences are 3.65, 2.01, 1.28 and 1.18 as derived from relative abundances, counts calibrated for differences in total number of 16S rRNA gene copies, SCML (by S. ruber ) and combined SCML (by S. ruber , A. acidiphilus and R. radiobacter ), respectively

    Techniques Used: Quantitative RT-PCR, Derivative Assay

    22) Product Images from "Modulation of integrin-linked kinase (ILK) expression in human oesophageal squamous cell carcinoma cell lines by the EGF and TGF?1 growth factors"

    Article Title: Modulation of integrin-linked kinase (ILK) expression in human oesophageal squamous cell carcinoma cell lines by the EGF and TGF?1 growth factors

    Journal: Cancer Cell International

    doi: 10.1186/1475-2867-6-12

    RT-PCR analysis of ILK expression in HOSCCs . RT-PCR was performed using specific primers co-amplifying ILK1 and ILK2. PCR products in the HOSCC cell lines WHCO1, WHCO3, WHCO5, WHCO6 and SNO separated on 2% agarose gels and stained with ethidium bromide. (A) An ILK band of 1360 bp was amplified across all five cell lines. 1 kb ladder in lane 1.
    Figure Legend Snippet: RT-PCR analysis of ILK expression in HOSCCs . RT-PCR was performed using specific primers co-amplifying ILK1 and ILK2. PCR products in the HOSCC cell lines WHCO1, WHCO3, WHCO5, WHCO6 and SNO separated on 2% agarose gels and stained with ethidium bromide. (A) An ILK band of 1360 bp was amplified across all five cell lines. 1 kb ladder in lane 1.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, Polymerase Chain Reaction, Staining, Amplification

    23) Product Images from "Karyotype Characterization of Nine Periwinkle Species (Gastropoda, Littorinidae)"

    Article Title: Karyotype Characterization of Nine Periwinkle Species (Gastropoda, Littorinidae)

    Journal: Genes

    doi: 10.3390/genes9110517

    Chromosomal mapping of rDNA and histone gene clusters in flat and common periwinkles. Fluorescent in situ hybridization mapping of minor rDNA (5S, red), H3 histone gene (H3, green) and major rDNA (28S, magenta) probes to mitotic chromosomes of the flat periwinkles Littorina obtusata and Littorina fabalis and the common periwinkle Littorina littorea counterstained with 4′-6-diamidino-2-phenylindole (DAPI). No minor (5S) rDNA signals were detected in the common periwinkle. Scale bars, 5 μm.
    Figure Legend Snippet: Chromosomal mapping of rDNA and histone gene clusters in flat and common periwinkles. Fluorescent in situ hybridization mapping of minor rDNA (5S, red), H3 histone gene (H3, green) and major rDNA (28S, magenta) probes to mitotic chromosomes of the flat periwinkles Littorina obtusata and Littorina fabalis and the common periwinkle Littorina littorea counterstained with 4′-6-diamidino-2-phenylindole (DAPI). No minor (5S) rDNA signals were detected in the common periwinkle. Scale bars, 5 μm.

    Techniques Used: In Situ Hybridization

    Chromosomal mapping of major rDNA and histone gene clusters in rough periwinkles. Fluorescent in situ hybridization mapping of H3 histone gene (H3, green) and major rDNA (28S, magenta) probes to mitotic chromosomes of the rough periwinkles Littorina arcana , Littorina saxatilis , and Littorina compressa counterstained with 4′-6-diamidino-2-phenylindole (DAPI). Scale bars, 5 μm.
    Figure Legend Snippet: Chromosomal mapping of major rDNA and histone gene clusters in rough periwinkles. Fluorescent in situ hybridization mapping of H3 histone gene (H3, green) and major rDNA (28S, magenta) probes to mitotic chromosomes of the rough periwinkles Littorina arcana , Littorina saxatilis , and Littorina compressa counterstained with 4′-6-diamidino-2-phenylindole (DAPI). Scale bars, 5 μm.

    Techniques Used: In Situ Hybridization

    24) Product Images from "Quality assessment of DNA derived from up to 30 years old formalin fixed paraffin embedded (FFPE) tissue for PCR-based methylation analysis using SMART-MSP and MS-HRM"

    Article Title: Quality assessment of DNA derived from up to 30 years old formalin fixed paraffin embedded (FFPE) tissue for PCR-based methylation analysis using SMART-MSP and MS-HRM

    Journal: BMC Cancer

    doi: 10.1186/1471-2407-9-453

    Melting profiles indicating that the DNA is heterogeneously methylated at the RARB promoter . If molecules, in which different CpG positions are methylated and unmethylated are amplified, the observed melting profile will be complex. These different molecules will form heteroduplexes and homoduplexes after the PCR and prior to the HRM step. The very early melting observed is likely to be the melting of various heteroduplexes. The intermediate and later melting are likely to be the melting of various homoduplexes with those having the highest number of CpG positions methylated melting the latest. The 100% methylated standards are indicated in red, the 10% methylated standards in blue, the 1% methylated standards in green, the 0.1% methylated standards in brown, the 0% methylated standard in orange and non template controls are indicated in black. A sample (in triplicate) judged to be heterogeneously methylated is indicated in turquoise.
    Figure Legend Snippet: Melting profiles indicating that the DNA is heterogeneously methylated at the RARB promoter . If molecules, in which different CpG positions are methylated and unmethylated are amplified, the observed melting profile will be complex. These different molecules will form heteroduplexes and homoduplexes after the PCR and prior to the HRM step. The very early melting observed is likely to be the melting of various heteroduplexes. The intermediate and later melting are likely to be the melting of various homoduplexes with those having the highest number of CpG positions methylated melting the latest. The 100% methylated standards are indicated in red, the 10% methylated standards in blue, the 1% methylated standards in green, the 0.1% methylated standards in brown, the 0% methylated standard in orange and non template controls are indicated in black. A sample (in triplicate) judged to be heterogeneously methylated is indicated in turquoise.

    Techniques Used: Methylation, Amplification, Polymerase Chain Reaction

    25) Product Images from "Quantitative Detection of Active Vibrios Associated with White Plague Disease in Mussismilia braziliensis Corals"

    Article Title: Quantitative Detection of Active Vibrios Associated with White Plague Disease in Mussismilia braziliensis Corals

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2017.02272

    Detection of Vibrio species by qPCR approach in coral samples from Abrolhos region. (A) Number of total Vibrio specie detected by 16S rRNA gene amplification. (B) Number of V. coralliilyticus detected by pyrH gene amplification. W, White Plague; H, healthy; BDL, below detection limit.
    Figure Legend Snippet: Detection of Vibrio species by qPCR approach in coral samples from Abrolhos region. (A) Number of total Vibrio specie detected by 16S rRNA gene amplification. (B) Number of V. coralliilyticus detected by pyrH gene amplification. W, White Plague; H, healthy; BDL, below detection limit.

    Techniques Used: Real-time Polymerase Chain Reaction, Amplification

    Amplification of target DNA in a background of closed related non-target. (A) Target and non-target DNA concentration used in the mixed reaction. (B) Copies detected by qPCR. All non-target DNA had 10-fold excess compare with target DNA. Target DNA i.e. (total Vibrio, V. harveyi , and V. coralliilyticus ) were diluted at least 1:100 while non-target (i.e., E. coli, V. campbellii, V. communis, V. tubiashii , and V. neptunius ) were diluted 1:10. Similarities of 16S rRNA between target and non-target are represented in line.
    Figure Legend Snippet: Amplification of target DNA in a background of closed related non-target. (A) Target and non-target DNA concentration used in the mixed reaction. (B) Copies detected by qPCR. All non-target DNA had 10-fold excess compare with target DNA. Target DNA i.e. (total Vibrio, V. harveyi , and V. coralliilyticus ) were diluted at least 1:100 while non-target (i.e., E. coli, V. campbellii, V. communis, V. tubiashii , and V. neptunius ) were diluted 1:10. Similarities of 16S rRNA between target and non-target are represented in line.

    Techniques Used: Amplification, Concentration Assay, Real-time Polymerase Chain Reaction

    26) Product Images from "TERT promoter mutations and gene amplification: Promoting TERT expression in Merkel cell carcinoma"

    Article Title: TERT promoter mutations and gene amplification: Promoting TERT expression in Merkel cell carcinoma

    Journal: Oncotarget

    doi:

    TERT mRNA expression and telomerase activity in MCC cell lines and tumors Relative TERT mRNA levels were expressed arbitrarily as the ratio of TERT and 18S rRNA (Frozen samples) or ACTB (FFPE samples) CT values. The level of telomerase activity was expressed arbitrarily as folds of that in HEK-293 cells. (A) TERT mRNA expression and (B) telomerase activity in MCC cell lines. (C) TERT mRNA levels in 33 FFPE MCC tumors. (D) TERT mRNA levels in 15 frozen MCC tumors and (E) telomerase activity in those same tumors. Patients MCCT_16b, _28, _30 and _33 had no materials available for telomerase activity assessment.
    Figure Legend Snippet: TERT mRNA expression and telomerase activity in MCC cell lines and tumors Relative TERT mRNA levels were expressed arbitrarily as the ratio of TERT and 18S rRNA (Frozen samples) or ACTB (FFPE samples) CT values. The level of telomerase activity was expressed arbitrarily as folds of that in HEK-293 cells. (A) TERT mRNA expression and (B) telomerase activity in MCC cell lines. (C) TERT mRNA levels in 33 FFPE MCC tumors. (D) TERT mRNA levels in 15 frozen MCC tumors and (E) telomerase activity in those same tumors. Patients MCCT_16b, _28, _30 and _33 had no materials available for telomerase activity assessment.

    Techniques Used: Expressing, Activity Assay, Formalin-fixed Paraffin-Embedded

    27) Product Images from "Formation of Linear Amplicons with Inverted Duplications in Leishmania Requires the MRE11 Nuclease"

    Article Title: Formation of Linear Amplicons with Inverted Duplications in Leishmania Requires the MRE11 Nuclease

    Journal: PLoS Genetics

    doi: 10.1371/journal.pgen.1004805

    Potential mechanisms for the formation of extrachromosomal linear amplicons. ( 1 ) Single-strand hairpin formation (a) or single-strand break (SSB) (near the IRs) during replication followed by annealing of the IRs (b), 3′-5′ exonuclease digestion of the exposed end (c) and DNA synthesis of the upstream region (d) and of the second strand to form an inverted duplication (g). ( 2 ) Double-strand break (DSB) near the IRs (e) followed by 3′-5′ exonuclease digestion at the DNA break of one strand (f), annealing of the IRs to form an inverted duplication (d) and synthesis of the second strand to generate linear amplicon (g). The new junction formed during annealing of the IRs can be detected by PCR using specific primers. IRs, inverted repeated sequences; ss, single-strand; SSB, single-strand break; DSB, double-strand break; p1–p2, primer pair used to detect the new junction. White rectangle: telomeric sequences.
    Figure Legend Snippet: Potential mechanisms for the formation of extrachromosomal linear amplicons. ( 1 ) Single-strand hairpin formation (a) or single-strand break (SSB) (near the IRs) during replication followed by annealing of the IRs (b), 3′-5′ exonuclease digestion of the exposed end (c) and DNA synthesis of the upstream region (d) and of the second strand to form an inverted duplication (g). ( 2 ) Double-strand break (DSB) near the IRs (e) followed by 3′-5′ exonuclease digestion at the DNA break of one strand (f), annealing of the IRs to form an inverted duplication (d) and synthesis of the second strand to generate linear amplicon (g). The new junction formed during annealing of the IRs can be detected by PCR using specific primers. IRs, inverted repeated sequences; ss, single-strand; SSB, single-strand break; DSB, double-strand break; p1–p2, primer pair used to detect the new junction. White rectangle: telomeric sequences.

    Techniques Used: DNA Synthesis, Amplification, Polymerase Chain Reaction

    28) Product Images from "Quantitative Detection of Active Vibrios Associated with White Plague Disease in Mussismilia braziliensis Corals"

    Article Title: Quantitative Detection of Active Vibrios Associated with White Plague Disease in Mussismilia braziliensis Corals

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2017.02272

    Detection of Vibrio species by qPCR approach in coral samples from Abrolhos region. (A) Number of total Vibrio specie detected by 16S rRNA gene amplification. (B) Number of V. coralliilyticus detected by pyrH gene amplification. W, White Plague; H, healthy; BDL, below detection limit.
    Figure Legend Snippet: Detection of Vibrio species by qPCR approach in coral samples from Abrolhos region. (A) Number of total Vibrio specie detected by 16S rRNA gene amplification. (B) Number of V. coralliilyticus detected by pyrH gene amplification. W, White Plague; H, healthy; BDL, below detection limit.

    Techniques Used: Real-time Polymerase Chain Reaction, Amplification

    Amplification of target DNA in a background of closed related non-target. (A) Target and non-target DNA concentration used in the mixed reaction. (B) Copies detected by qPCR. All non-target DNA had 10-fold excess compare with target DNA. Target DNA i.e. (total Vibrio, V. harveyi , and V. coralliilyticus ) were diluted at least 1:100 while non-target (i.e., E. coli, V. campbellii, V. communis, V. tubiashii , and V. neptunius ) were diluted 1:10. Similarities of 16S rRNA between target and non-target are represented in line.
    Figure Legend Snippet: Amplification of target DNA in a background of closed related non-target. (A) Target and non-target DNA concentration used in the mixed reaction. (B) Copies detected by qPCR. All non-target DNA had 10-fold excess compare with target DNA. Target DNA i.e. (total Vibrio, V. harveyi , and V. coralliilyticus ) were diluted at least 1:100 while non-target (i.e., E. coli, V. campbellii, V. communis, V. tubiashii , and V. neptunius ) were diluted 1:10. Similarities of 16S rRNA between target and non-target are represented in line.

    Techniques Used: Amplification, Concentration Assay, Real-time Polymerase Chain Reaction

    29) Product Images from "N-glycan alterations are associated with drug resistance in human hepatocellular carcinoma"

    Article Title: N-glycan alterations are associated with drug resistance in human hepatocellular carcinoma

    Journal: Molecular Cancer

    doi: 10.1186/1476-4598-6-32

    RT-PCR analysis of glycosyltransferase mRNA expression in each cell line . Detailed experimental procedures are in Materials and Methods. Human glyceraldehyde-3-phosphate dehydrogenase (hGAPDH) is reported for internal controls.
    Figure Legend Snippet: RT-PCR analysis of glycosyltransferase mRNA expression in each cell line . Detailed experimental procedures are in Materials and Methods. Human glyceraldehyde-3-phosphate dehydrogenase (hGAPDH) is reported for internal controls.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing

    30) Product Images from "Actinomycin D Induces High-Level Resistance to Thymidine Analogs in Replication of Human Immunodeficiency Virus Type 1 by Interfering with Host Cell Thymidine Kinase Expression"

    Article Title: Actinomycin D Induces High-Level Resistance to Thymidine Analogs in Replication of Human Immunodeficiency Virus Type 1 by Interfering with Host Cell Thymidine Kinase Expression

    Journal: Journal of Virology

    doi: 10.1128/JVI.77.2.1011-1020.2003

    ActD treatment downregulates transcription of TK-1. MT-2 cells were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 4 days in the absence or presence of 10 nM ActD. (A) Cells were washed and RNA was isolated. The RNA was treated with DNase and subsequently used to synthesize cDNA; the cDNA (1 μg) was serially diluted 1:4. The diluted cDNA served as template for subsequent PCRs to amplify TK-1 and β-actin. To confirm the absence of contaminating DNA, control cDNA reactions were prepared in which RT was omitted from each reaction. Amplified products were not seen in these controls (data not shown). β-Actin served as an internal control to monitor the efficiency of amplification. The PCR product sizes of TK-1 and β-actin were 540 and 353 bp, respectively. MW refers to a molecular marker consisting of a 100-bp ladder (Invitrogen). (B) The infected cells were washed and cellular protein was extracted using RIPA buffer (see Materials and Methods). For each blot, 50 μg of total cellular protein was loaded on a 12% bis-Tris sodium dodecyl sulfate-acrylamide gel, and Western blotting was performed using an anti-TK-1 antibody. Subsequently, the antibody was stripped and then the blot was reprobed using anti-actin antibody. Images were quantitated using the Epi chemi II Darkroom and LabWork software (Ultra-violet Products, Ltd., Upland, Calif.).
    Figure Legend Snippet: ActD treatment downregulates transcription of TK-1. MT-2 cells were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 4 days in the absence or presence of 10 nM ActD. (A) Cells were washed and RNA was isolated. The RNA was treated with DNase and subsequently used to synthesize cDNA; the cDNA (1 μg) was serially diluted 1:4. The diluted cDNA served as template for subsequent PCRs to amplify TK-1 and β-actin. To confirm the absence of contaminating DNA, control cDNA reactions were prepared in which RT was omitted from each reaction. Amplified products were not seen in these controls (data not shown). β-Actin served as an internal control to monitor the efficiency of amplification. The PCR product sizes of TK-1 and β-actin were 540 and 353 bp, respectively. MW refers to a molecular marker consisting of a 100-bp ladder (Invitrogen). (B) The infected cells were washed and cellular protein was extracted using RIPA buffer (see Materials and Methods). For each blot, 50 μg of total cellular protein was loaded on a 12% bis-Tris sodium dodecyl sulfate-acrylamide gel, and Western blotting was performed using an anti-TK-1 antibody. Subsequently, the antibody was stripped and then the blot was reprobed using anti-actin antibody. Images were quantitated using the Epi chemi II Darkroom and LabWork software (Ultra-violet Products, Ltd., Upland, Calif.).

    Techniques Used: Infection, Cell Culture, Isolation, Amplification, Polymerase Chain Reaction, Marker, Acrylamide Gel Assay, Western Blot, Software

    ActD upregulates HIV-1 replication but not HIV-2 replication. (A) MT-2 (closed circles), MT-4 (open circles), Jurkat (open triangles), or PHA-stimulated PBMCs (closed squares) were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 7 days in the presence of various concentrations of ActD (0 to 1,000 nM). HIV-1 replication was measured by a p24 antigen capture assay. Results show the percentage of virus growth in ActD-treated cells compared to that in untreated cells. In this experiment, the p24 concentrations in the culture supernatant of MT-2, MT-4, Jurkat, and PBMCs in the absence of ActD were 159 ± 33, 56 ± 11, 9.1 ± 2.1, and 10 ± 1.4 ng/ml, respectively. (B) HIV-1 NL4.3 -infected MT-2 cells were cultured in 10 ml of RPMI-10 in T-25 cm 2 culture flasks in the presence (closed circles) or absence (open circles) of 10 nM ActD, and culture supernatants were collected every day. The assay was performed in triplicate, and the p24 levels in culture supernatants were measured by the p24 antigen capture assay. (C) MT-2 cells were infected with HIV-1 NL4.3 (white bar), HIV-1 DH12 (gray bar), or HIV-2 MVP15132 (black bar) for 2 h at 37°C. After washing, the MT-2 cells were cultured for an additional 7 days at 37°C in the presence of 0 to 100 nM ActD. The p24 (for HIV-1) or p27 (for HIV-2) antigen levels in the culture supernatants were measured by p24 or p27 antigen capture assays. (D) MT-2 cells were infected with HIV-1 NL4.3 and cultured for 4 days in the absence or presence of 10 nM ActD.
    Figure Legend Snippet: ActD upregulates HIV-1 replication but not HIV-2 replication. (A) MT-2 (closed circles), MT-4 (open circles), Jurkat (open triangles), or PHA-stimulated PBMCs (closed squares) were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 7 days in the presence of various concentrations of ActD (0 to 1,000 nM). HIV-1 replication was measured by a p24 antigen capture assay. Results show the percentage of virus growth in ActD-treated cells compared to that in untreated cells. In this experiment, the p24 concentrations in the culture supernatant of MT-2, MT-4, Jurkat, and PBMCs in the absence of ActD were 159 ± 33, 56 ± 11, 9.1 ± 2.1, and 10 ± 1.4 ng/ml, respectively. (B) HIV-1 NL4.3 -infected MT-2 cells were cultured in 10 ml of RPMI-10 in T-25 cm 2 culture flasks in the presence (closed circles) or absence (open circles) of 10 nM ActD, and culture supernatants were collected every day. The assay was performed in triplicate, and the p24 levels in culture supernatants were measured by the p24 antigen capture assay. (C) MT-2 cells were infected with HIV-1 NL4.3 (white bar), HIV-1 DH12 (gray bar), or HIV-2 MVP15132 (black bar) for 2 h at 37°C. After washing, the MT-2 cells were cultured for an additional 7 days at 37°C in the presence of 0 to 100 nM ActD. The p24 (for HIV-1) or p27 (for HIV-2) antigen levels in the culture supernatants were measured by p24 or p27 antigen capture assays. (D) MT-2 cells were infected with HIV-1 NL4.3 and cultured for 4 days in the absence or presence of 10 nM ActD.

    Techniques Used: Infection, Cell Culture

    ActD treatment reverses the inhibition of HIV replication by RT inhibitors. MT-2 cells were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 7 days in the presence of various concentrations of ActD (0 to 100 nM) without AZT (open circles) or with 1 μM AZT (closed circles). HIV-1 replication was measured by a p24 antigen capture assay.
    Figure Legend Snippet: ActD treatment reverses the inhibition of HIV replication by RT inhibitors. MT-2 cells were infected with HIV-1 NL4.3 for 2 h at 37°C. The infected cells were cultured for 7 days in the presence of various concentrations of ActD (0 to 100 nM) without AZT (open circles) or with 1 μM AZT (closed circles). HIV-1 replication was measured by a p24 antigen capture assay.

    Techniques Used: Inhibition, Infection, Cell Culture

    31) Product Images from "IL-12p40 Homodimer Ameliorates Experimental Autoimmune Arthritis"

    Article Title: IL-12p40 Homodimer Ameliorates Experimental Autoimmune Arthritis

    Journal: The Journal of Immunology Author Choice

    doi: 10.4049/jimmunol.1500400

    (p40) 2 induces CD4 + CD25 + Foxp + Tregs in vivo and in vitro. ( A and B ) Spleen and joint tissue from (p40) 2 -injected CIA and control mice were stained with anti-mouse CD4-PE, anti-mouse CD25-allophycocyanin, and anti-mouse Foxp3-FITC. Stained spleen tissue was analyzed using a confocal microscope (original magnification ×400). Arrowheads indicate Treg or Th17 cells. Tregs are purple. Data shown are representative of three independent experiments. (B and C ) Spleen cells were isolated from CIA mice. The cells were cultured with IL-23 (10 ng/ml) and IL-23 plus (p40) 2 (10 ng/ml) for 3 d. (B) Foxp3 protein was measured in cell lysates by Western blot analysis using the Foxp3-specific Ab. (C) Cultured cells were stained with anti-mouse CD4-PerCP, anti-mouse CD25-FITC, and anti-mouse Foxp3-PE. CD4 + CD25 + Foxp3 + Tregs were analyzed using FlowJo software. ( D ) RORγt and Foxp3 mRNA expression was measured in spleen cells by real-time PCR. ( E ) IL-17, TGF-β, and IL-10 mRNA expression was measured in spleen cells by real-time PCR. Data are mean ± SD are representative of three independent experiments. * p
    Figure Legend Snippet: (p40) 2 induces CD4 + CD25 + Foxp + Tregs in vivo and in vitro. ( A and B ) Spleen and joint tissue from (p40) 2 -injected CIA and control mice were stained with anti-mouse CD4-PE, anti-mouse CD25-allophycocyanin, and anti-mouse Foxp3-FITC. Stained spleen tissue was analyzed using a confocal microscope (original magnification ×400). Arrowheads indicate Treg or Th17 cells. Tregs are purple. Data shown are representative of three independent experiments. (B and C ) Spleen cells were isolated from CIA mice. The cells were cultured with IL-23 (10 ng/ml) and IL-23 plus (p40) 2 (10 ng/ml) for 3 d. (B) Foxp3 protein was measured in cell lysates by Western blot analysis using the Foxp3-specific Ab. (C) Cultured cells were stained with anti-mouse CD4-PerCP, anti-mouse CD25-FITC, and anti-mouse Foxp3-PE. CD4 + CD25 + Foxp3 + Tregs were analyzed using FlowJo software. ( D ) RORγt and Foxp3 mRNA expression was measured in spleen cells by real-time PCR. ( E ) IL-17, TGF-β, and IL-10 mRNA expression was measured in spleen cells by real-time PCR. Data are mean ± SD are representative of three independent experiments. * p

    Techniques Used: In Vivo, In Vitro, Injection, Mouse Assay, Staining, Microscopy, Isolation, Cell Culture, Western Blot, Software, Expressing, Real-time Polymerase Chain Reaction

    32) Product Images from "SMAC mimetics induce autophagy-dependent apoptosis of HIV-1-infected resting memory CD4+ T cells"

    Article Title: SMAC mimetics induce autophagy-dependent apoptosis of HIV-1-infected resting memory CD4+ T cells

    Journal: Cell host & microbe

    doi: 10.1016/j.chom.2018.09.007

    SMAC mimetics preferentially induce cell death in HIV-T CM . ( A ) T CM and HIV-T CM were treated with SM or 1 μ . DNA was extracted from HIV-T CM . n = 4. ( B ) ELISA performed for HIV p24 antigen in supernatants from cells treated in A . n = 4. ( C ) RT-qPCR performed for extracellular release of HIV gag mRNA from cells treated in A . n = 4. ( D ) T CM and HIV-T CM were treated with SM for 24 h. Top , representative western blots of PARP1 and CASP8 cleavage (cPARP1 and cCASP8) using antibody to PARP1, CASP8, and ACTB. Bottom , densitometric analysis of blots, n = 4. ( E ) T CM and HIV-T CM . n = 4. ( F ) HIV-T CM were pretreated with vehicle control or TNF neutralizing antibody 2 h before incubation with SM for 24 h. Cell death was measured using a cell death ELISA. n = 4. ( G ) Resting CD4+ T cells were isolated from HIV infected donors on suppressive antiretroviral therapy, viral load
    Figure Legend Snippet: SMAC mimetics preferentially induce cell death in HIV-T CM . ( A ) T CM and HIV-T CM were treated with SM or 1 μ . DNA was extracted from HIV-T CM . n = 4. ( B ) ELISA performed for HIV p24 antigen in supernatants from cells treated in A . n = 4. ( C ) RT-qPCR performed for extracellular release of HIV gag mRNA from cells treated in A . n = 4. ( D ) T CM and HIV-T CM were treated with SM for 24 h. Top , representative western blots of PARP1 and CASP8 cleavage (cPARP1 and cCASP8) using antibody to PARP1, CASP8, and ACTB. Bottom , densitometric analysis of blots, n = 4. ( E ) T CM and HIV-T CM . n = 4. ( F ) HIV-T CM were pretreated with vehicle control or TNF neutralizing antibody 2 h before incubation with SM for 24 h. Cell death was measured using a cell death ELISA. n = 4. ( G ) Resting CD4+ T cells were isolated from HIV infected donors on suppressive antiretroviral therapy, viral load

    Techniques Used: Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Western Blot, Incubation, Isolation, Infection

    33) Product Images from "Diagnostic value of the molecular detection of Sarcoptes scabiei from a skin scraping in patients with suspected scabies"

    Article Title: Diagnostic value of the molecular detection of Sarcoptes scabiei from a skin scraping in patients with suspected scabies

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0008229

    Detection of Sarcoptes scabiei DNA by polymerase chain reaction (PCR). DNA was purified from skin scraping samples from suspected scabies patients (lanes 1–5), and run by electrophoresis with a positive control (lane 6), and a negative control (lane 7). Lane M is a 100-bp DNA ladder marker.
    Figure Legend Snippet: Detection of Sarcoptes scabiei DNA by polymerase chain reaction (PCR). DNA was purified from skin scraping samples from suspected scabies patients (lanes 1–5), and run by electrophoresis with a positive control (lane 6), and a negative control (lane 7). Lane M is a 100-bp DNA ladder marker.

    Techniques Used: Polymerase Chain Reaction, Purification, Electrophoresis, Positive Control, Negative Control, Marker

    34) Product Images from "Upregulation of microRNA-4417 and Its Target Genes Contribute to Nickel Chloride-promoted Lung Epithelial Cell Fibrogenesis and Tumorigenesis"

    Article Title: Upregulation of microRNA-4417 and Its Target Genes Contribute to Nickel Chloride-promoted Lung Epithelial Cell Fibrogenesis and Tumorigenesis

    Journal: Scientific Reports

    doi: 10.1038/s41598-017-14610-7

    NiCl 2 correlates with miRNA expression patterns. ( a ) BEAS-2B cells (1 × 10 6 cells/6 cm dish) were treated with NiCl 2 (0.5 mM) for 48 h. Heat maps and cluster analysis of the 46 miRNA differentially expressed between control and 0.5 mM NiCl 2 -treated BEAS-2B cells. Up- and down-regulated genes are represented in red and green, respectively. ( b ) Ten of the 46 miRNAs identified on microarray were corroborated with qRT-PCR data. BEAS-2B cells were treated with NiCl 2 (0, 0.5 mM) for 48 h. The log2 of the ratios of expression levels are shown. ( c ) Seven miRNAs identified as significantly different between Ni-treated A549 cells and controls on microarray study were evaluated on qRT-PCR. All values have been normalized to the level of RNU6B and are the averages of three independent readings. *p
    Figure Legend Snippet: NiCl 2 correlates with miRNA expression patterns. ( a ) BEAS-2B cells (1 × 10 6 cells/6 cm dish) were treated with NiCl 2 (0.5 mM) for 48 h. Heat maps and cluster analysis of the 46 miRNA differentially expressed between control and 0.5 mM NiCl 2 -treated BEAS-2B cells. Up- and down-regulated genes are represented in red and green, respectively. ( b ) Ten of the 46 miRNAs identified on microarray were corroborated with qRT-PCR data. BEAS-2B cells were treated with NiCl 2 (0, 0.5 mM) for 48 h. The log2 of the ratios of expression levels are shown. ( c ) Seven miRNAs identified as significantly different between Ni-treated A549 cells and controls on microarray study were evaluated on qRT-PCR. All values have been normalized to the level of RNU6B and are the averages of three independent readings. *p

    Techniques Used: Expressing, Microarray, Quantitative RT-PCR

    Network of miRNA target genes in nickel-treated cells. ( a ) Octagonal shapes represent mRNAs and diamond shape represents miR-4417. The figure was generated using CytoScape®. ( b ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 48 h. The mRNA levels were analyzed on RT-PCR. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, NAP1L5/β-actin and TAB2/β-actin are shown. ( c ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 72 h and the protein levels were determined on Western blot. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, RAB6A/β-actin, p-TAB2/β-actin and TAB2/β-actin are shown.
    Figure Legend Snippet: Network of miRNA target genes in nickel-treated cells. ( a ) Octagonal shapes represent mRNAs and diamond shape represents miR-4417. The figure was generated using CytoScape®. ( b ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 48 h. The mRNA levels were analyzed on RT-PCR. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, NAP1L5/β-actin and TAB2/β-actin are shown. ( c ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 72 h and the protein levels were determined on Western blot. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, RAB6A/β-actin, p-TAB2/β-actin and TAB2/β-actin are shown.

    Techniques Used: Generated, Reverse Transcription Polymerase Chain Reaction, Western Blot

    35) Product Images from "Direct Detection and Differentiation of Legionella spp. and Legionella pneumophila in Clinical Specimens by Dual-Color Real-Time PCR and Melting Curve Analysis"

    Article Title: Direct Detection and Differentiation of Legionella spp. and Legionella pneumophila in Clinical Specimens by Dual-Color Real-Time PCR and Melting Curve Analysis

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.40.10.3814-3817.2002

    Evaluation of the dual-color Legionella PCR assay with clinical specimens. A representative set of 23 L. pneumophila culture-positive BAL specimens, 4 Legionella culture-negative BAL specimens, and 3 Legionella culture-negative BAL specimens spiked with cultured L. longbeachae , L. micdadei , or L. bozemanii organisms was analyzed. Ten nanograms of L. pneumophila serogroup 1 (ATCC 33152) template DNA was used as a positive control. (A) Fluorescence readout at 640 nm showing the LightCycler results with the Legionella spp.-specific set of hybridization probes Leg-HP-1 and Leg-HP-2. (B) Fluorescence readout at 705 nm showing the LightCycler results with the L. pneumophila -specific set of hybridization probes Lpn-HP-1 and Lpn-HP-2.
    Figure Legend Snippet: Evaluation of the dual-color Legionella PCR assay with clinical specimens. A representative set of 23 L. pneumophila culture-positive BAL specimens, 4 Legionella culture-negative BAL specimens, and 3 Legionella culture-negative BAL specimens spiked with cultured L. longbeachae , L. micdadei , or L. bozemanii organisms was analyzed. Ten nanograms of L. pneumophila serogroup 1 (ATCC 33152) template DNA was used as a positive control. (A) Fluorescence readout at 640 nm showing the LightCycler results with the Legionella spp.-specific set of hybridization probes Leg-HP-1 and Leg-HP-2. (B) Fluorescence readout at 705 nm showing the LightCycler results with the L. pneumophila -specific set of hybridization probes Lpn-HP-1 and Lpn-HP-2.

    Techniques Used: Polymerase Chain Reaction, Cell Culture, Positive Control, Fluorescence, Hybridization

    36) Product Images from "GC Content-Based Pan-Pox Universal PCR Assays for Poxvirus Detection ▿"

    Article Title: GC Content-Based Pan-Pox Universal PCR Assays for Poxvirus Detection ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.01697-09

    Low- and high-GC PCR and TaqI RFLP assays. (A) Low-GC PCR amplicons and TaqI RFLP patterns. The top panel shows low-GC PCR amplicons from 20 poxvirus DNA samples. The bottom panel shows corresponding TaqI restriction endonuclease RFLP patterns. (B) High-GC
    Figure Legend Snippet: Low- and high-GC PCR and TaqI RFLP assays. (A) Low-GC PCR amplicons and TaqI RFLP patterns. The top panel shows low-GC PCR amplicons from 20 poxvirus DNA samples. The bottom panel shows corresponding TaqI restriction endonuclease RFLP patterns. (B) High-GC

    Techniques Used: Polymerase Chain Reaction

    37) Product Images from "Local overexpression of the myostatin propeptide increases glucose transporter expression and enhances skeletal muscle glucose disposal"

    Article Title: Local overexpression of the myostatin propeptide increases glucose transporter expression and enhances skeletal muscle glucose disposal

    Journal: American Journal of Physiology - Endocrinology and Metabolism

    doi: 10.1152/ajpendo.00586.2013

    Effect of myostatin blockade on selected mediators of muscle size and metabolism. Total RNA and cDNA was prepared from paired TC muscles, and SYBR Green Real-time PCR was conducted using 20 ng of each cDNA or DNA standard in duplicate and primers as listed in materials and methods . Summary data (means ± SE; n = 10–12) are shown for each target mRNA of interest [insulin-like growth factor-I (IGF-I), IGF-I receptor (IGF-I Rec), FOXO3, peroxisome proliferator-activated receptor coactivator 1α (PGC1a), nuclear factor-κB p65 subunit (p65), activin IIB receptor (activin 2BR), latent transforming growth factor-β-binding protein (LTBP3), muscle ring finger protein 1 (MURF1), and atrogin 1], normalized to the geometric mean of the reference mRNAs cyclophilin, 36B4, and 18S. Filled bars, control; open bars, test muscle. * P
    Figure Legend Snippet: Effect of myostatin blockade on selected mediators of muscle size and metabolism. Total RNA and cDNA was prepared from paired TC muscles, and SYBR Green Real-time PCR was conducted using 20 ng of each cDNA or DNA standard in duplicate and primers as listed in materials and methods . Summary data (means ± SE; n = 10–12) are shown for each target mRNA of interest [insulin-like growth factor-I (IGF-I), IGF-I receptor (IGF-I Rec), FOXO3, peroxisome proliferator-activated receptor coactivator 1α (PGC1a), nuclear factor-κB p65 subunit (p65), activin IIB receptor (activin 2BR), latent transforming growth factor-β-binding protein (LTBP3), muscle ring finger protein 1 (MURF1), and atrogin 1], normalized to the geometric mean of the reference mRNAs cyclophilin, 36B4, and 18S. Filled bars, control; open bars, test muscle. * P

    Techniques Used: SYBR Green Assay, Real-time Polymerase Chain Reaction, Binding Assay

    38) Product Images from "Upregulation of microRNA-4417 and Its Target Genes Contribute to Nickel Chloride-promoted Lung Epithelial Cell Fibrogenesis and Tumorigenesis"

    Article Title: Upregulation of microRNA-4417 and Its Target Genes Contribute to Nickel Chloride-promoted Lung Epithelial Cell Fibrogenesis and Tumorigenesis

    Journal: Scientific Reports

    doi: 10.1038/s41598-017-14610-7

    NiCl 2 correlates with miRNA expression patterns. ( a ) BEAS-2B cells (1 × 10 6 cells/6 cm dish) were treated with NiCl 2 (0.5 mM) for 48 h. Heat maps and cluster analysis of the 46 miRNA differentially expressed between control and 0.5 mM NiCl 2 -treated BEAS-2B cells. Up- and down-regulated genes are represented in red and green, respectively. ( b ) Ten of the 46 miRNAs identified on microarray were corroborated with qRT-PCR data. BEAS-2B cells were treated with NiCl 2 (0, 0.5 mM) for 48 h. The log2 of the ratios of expression levels are shown. ( c ) Seven miRNAs identified as significantly different between Ni-treated A549 cells and controls on microarray study were evaluated on qRT-PCR. All values have been normalized to the level of RNU6B and are the averages of three independent readings. *p
    Figure Legend Snippet: NiCl 2 correlates with miRNA expression patterns. ( a ) BEAS-2B cells (1 × 10 6 cells/6 cm dish) were treated with NiCl 2 (0.5 mM) for 48 h. Heat maps and cluster analysis of the 46 miRNA differentially expressed between control and 0.5 mM NiCl 2 -treated BEAS-2B cells. Up- and down-regulated genes are represented in red and green, respectively. ( b ) Ten of the 46 miRNAs identified on microarray were corroborated with qRT-PCR data. BEAS-2B cells were treated with NiCl 2 (0, 0.5 mM) for 48 h. The log2 of the ratios of expression levels are shown. ( c ) Seven miRNAs identified as significantly different between Ni-treated A549 cells and controls on microarray study were evaluated on qRT-PCR. All values have been normalized to the level of RNU6B and are the averages of three independent readings. *p

    Techniques Used: Expressing, Microarray, Quantitative RT-PCR

    Network of miRNA target genes in nickel-treated cells. ( a ) Octagonal shapes represent mRNAs and diamond shape represents miR-4417. The figure was generated using CytoScape®. ( b ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 48 h. The mRNA levels were analyzed on RT-PCR. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, NAP1L5/β-actin and TAB2/β-actin are shown. ( c ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 72 h and the protein levels were determined on Western blot. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, RAB6A/β-actin, p-TAB2/β-actin and TAB2/β-actin are shown.
    Figure Legend Snippet: Network of miRNA target genes in nickel-treated cells. ( a ) Octagonal shapes represent mRNAs and diamond shape represents miR-4417. The figure was generated using CytoScape®. ( b ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 48 h. The mRNA levels were analyzed on RT-PCR. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, NAP1L5/β-actin and TAB2/β-actin are shown. ( c ) BEAS-2B (1 × 10 6 cells/6 cm dish) and A549 (5 × 10 5 cells/6 cm dish) cells were treated with NiCl 2 (0, 0.25, 0.5 mM and 0, 0.5, 1 mM, respectively) for 72 h and the protein levels were determined on Western blot. β-actin was used as the internal control. The relative ratios of ENOSF1/β-actin, RAB6A/β-actin, p-TAB2/β-actin and TAB2/β-actin are shown.

    Techniques Used: Generated, Reverse Transcription Polymerase Chain Reaction, Western Blot

    39) Product Images from "Repair and Polyadenylation of a Naturally Occurring Hepatitis C Virus 3? Nontranslated Region-Shorter Variant in Selectable Replicon Cell Lines"

    Article Title: Repair and Polyadenylation of a Naturally Occurring Hepatitis C Virus 3? Nontranslated Region-Shorter Variant in Selectable Replicon Cell Lines

    Journal: Journal of Virology

    doi: 10.1128/JVI.80.9.4336-4343.2006

    ; the present study). (C) Effect of 3′-end deletions on cell colony formation using selectable replicons. WT is the pFK5.1 (Con1) construct amplified using T7 primer and reverse primer corresponding to the 3′ end (see the text for further details). Numbers below the plates refer to the CFU per microgram of in vitro-transcribed replicon RNA.
    Figure Legend Snippet: ; the present study). (C) Effect of 3′-end deletions on cell colony formation using selectable replicons. WT is the pFK5.1 (Con1) construct amplified using T7 primer and reverse primer corresponding to the 3′ end (see the text for further details). Numbers below the plates refer to the CFU per microgram of in vitro-transcribed replicon RNA.

    Techniques Used: Construct, Amplification, In Vitro

    Colony formation of selectable replicons carrying a poly(A) tail. The top images show the secondary structures of 3′ SL1 structures including the poly(A) tail; the bottom images show colonies formed using replicons containing poly(A) tails (pFK5.1_pA57 and pFK5.1_pA65) and the WT 3′ end (pFK5.1C -5 U). The numbers below the plates refer to the CFU per microgram of in vitro-transcribed replicon RNA.
    Figure Legend Snippet: Colony formation of selectable replicons carrying a poly(A) tail. The top images show the secondary structures of 3′ SL1 structures including the poly(A) tail; the bottom images show colonies formed using replicons containing poly(A) tails (pFK5.1_pA57 and pFK5.1_pA65) and the WT 3′ end (pFK5.1C -5 U). The numbers below the plates refer to the CFU per microgram of in vitro-transcribed replicon RNA.

    Techniques Used: In Vitro

    40) Product Images from "Transmission of H7N9 Influenza Viruses with a Polymorphism at PB2 Residue 627 in Chickens and Ferrets"

    Article Title: Transmission of H7N9 Influenza Viruses with a Polymorphism at PB2 Residue 627 in Chickens and Ferrets

    Journal: Journal of Virology

    doi: 10.1128/JVI.01444-15

    Transmission of the SCk1772 virus in Re-6-vaccinated chickens. Chickens were vaccinated with Re-6 H5N1 vaccine. Donor chickens were inoculated with 5 × 10 6 EID 50 s of SCk1772, and naive direct-contact chickens were introduced at 1 dpi. Oropharyngeal swabs (A) and cloacal swabs (B) were quantified by H7 qRRT-PCR. The mean viral RNA copy ± SEM is shown, with the solid line representing the limit of detection at 3.14 copies/μl. The proportion of chickens that seroconverted is shown above each group.
    Figure Legend Snippet: Transmission of the SCk1772 virus in Re-6-vaccinated chickens. Chickens were vaccinated with Re-6 H5N1 vaccine. Donor chickens were inoculated with 5 × 10 6 EID 50 s of SCk1772, and naive direct-contact chickens were introduced at 1 dpi. Oropharyngeal swabs (A) and cloacal swabs (B) were quantified by H7 qRRT-PCR. The mean viral RNA copy ± SEM is shown, with the solid line representing the limit of detection at 3.14 copies/μl. The proportion of chickens that seroconverted is shown above each group.

    Techniques Used: Transmission Assay, Polymerase Chain Reaction

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    Article Snippet: .. Extraction of DNA and RNA from Cells DNA and RNA were extracted from transfected and non-transfected hepatoma cells in a Magnapure robot (Roche Applied Science, Germany) using the Total NA protocol. .. Harvested cells were washed in 1 mL PBS and after centrifugation at 5000 rpm for 3 min the pellet was re-suspended in 800 µL RLT lysis buffer (Qiagen Sciences, MD, USA) before extraction.

    Synthesized:

    Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
    Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

    Isolation:

    Article Title: MERS-coronavirus replication induces severe in vitro cytopathology and is strongly inhibited by cyclosporin A or interferon-? treatment
    Article Snippet: .. RNA from 200 µl culture medium of CoV-infected cells was isolated with a MagnaPure LC Total Nucleic Acid Isolation kit (Roche) and eluted in 100 µl. .. RT-PCR conditions for quantifying MERS-CoV and SARS-CoV RNA and amplification parameters have been described previously ( ; ).

    Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
    Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

    Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
    Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

    Quantitative RT-PCR:

    Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
    Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

    Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
    Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

    Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
    Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

    Purification:

    Article Title: Homologous Recombination Occurs in Entamoeba and Is Enhanced during Growth Stress and Stage Conversion
    Article Snippet: .. RNA purification and Real Time RT-PCR Entamoeba cells were harvested and suspended immediately in TriZol reagent (Invitrogen) and RNA was isolated and treated with DNase I (Roche) according to manufacturer's protocol. .. RNA was reverse transcribed into cDNA using Superscript III reverse transcriptase (Invitrogen) and random hexamers used as a primer.

    Real-time Polymerase Chain Reaction:

    Article Title: Genome-wide analyses reveal the IRE1a-XBP1 pathway promotes T helper cell differentiation by resolving secretory stress and accelerating proliferation
    Article Snippet: .. Reverse transcription quantitative PCR (RT-qPCR) Total RNA was isolated from two million cells by SV total RNA isolation kit (Promega). cDNA was prepared by annealing 500 ng RNA with oligo dT as per the manufacturer’s instructions (Transcriptor High Fidelity cDNA Synthesis kit, Roche). ..

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1, et al. Chronic hypoxia‐induced slug promotes invasive behavior of prostate cancer cells by activating expression of ephrin‐B1
    Article Snippet: .. 2.5 Real‐time quantitative RT‐PCR First‐strand cDNA was synthesized from the total RNA using ThermoScript RT‐PCR System (Roche, Indianapolis, IN, USA). .. PCR was performed on a LightCycler system (Roche) using LightCycler FastStart DNA Master SYBR Green I reaction mix (Roche) and QuantiTect Primer Assays (QIAGEN, Hilden, Germany).

    Article Title: Arabidopsis ETO1 specifically interacts with and negatively regulates type 2 1-aminocyclopropane-1-carboxylate synthases
    Article Snippet: .. RNA (1 μg) was used in each RT-PCR following the manufacturer's protocol (Titan One Tube RT-PCR System, Roche Diagnostics). ..

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  • 90
    Roche polymerase chain reaction pcr mixture
    Polymerase Chain Reaction Pcr Mixture, supplied by Roche, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/polymerase chain reaction pcr mixture/product/Roche
    Average 90 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    polymerase chain reaction pcr mixture - by Bioz Stars, 2020-09
    90/100 stars
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