dna sequences Search Results


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  • 99
    Thermo Fisher 3730xl dna analyzer
    3730xl Dna Analyzer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 46651 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore dna sequence
    Generation of a <t>bb0028</t> mutant. A . An IPTG-regulatable bb0028 mutant was generated by inserting a streptomycin resistance cassette followed by the flacp promoter immediately upstream of bb0028 . Red arrows indicate primer-binding areas used to confirm insertion of the streptomycin cassette and flacp promoter into the borrelial genome. B . PCR amplification resulted in a 1.2 kb amplicon in the wildtype strain (WT), as expected, and a 3.0 kb amplicon in the BB0028 mutant (flacp::0028) indicating the streptomycin cassette and flacp promoter were inserted in the mutant. A PCR reaction with no <t>DNA</t> template (Neg) was included as a negative control. DNA sizes, in kilobase pairs, are indicated at left. C . Whole-cell lysate from the BB0028 mutant (flacp::0028) strain propagated with (+) or without (−) 1 mM IPTG, as well as from the parental, wildtype strain (WT), were subjected to immunoblot analysis using BB0028 specific antibodies (top panel) or FlaB antibodies (bottom panel). FlaB reactivity was used to ensure all lanes were loaded equally.
    Dna Sequence, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1652 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Illumina Inc sequencing rna
    Validation of gene expression changes estimated with CANEapp with quantitative real-time PCR. a <t>RNA-seq</t> analysis of hippocampi of Alzheimer’s disease patients and controls. Hippocampal tissue from 4 AD patients and 4 control individuals was used to extract total RNA and perform ribodepletion and strand-specific library preparation. Single-end RNA sequencing was performed on <t>Illumina</t> HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes measured with real-time PCR was compared with expression values generated by CANEapp. b RNA-seq of developing mouse cortex. Tissue from 4 embryonic day 17 and 3 adult mouse cortical samples was processed to extract polyA-selected RNA and generate paired-end unidirectional sequencing data with Illumina Genome Analyzer IIx. Gene expression estimates of 4 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. c Fold changes of gene expression for RNA-seq of liver of rats treated with two DNA-damage compounds. The data was produced by paired-end sequencing of polyA-selected RNA on Illumina HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. R 2 -coefficient of determination
    Sequencing Rna, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 92/100, based on 685 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Biotechnology Information dna sequences
    Agarose gels of <t>amplicons</t> generated by PCR amplification of enriched fecal specimens using the invE-A (457-bp amplicon) and hisJ (496-bp amplicon) primer sets. (A) Results using the invE-A primers. Lanes 1 and 6 contain molecular weight standards (100-bp <t>DNA</t> ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lane 3, DNA extracted from culture-negative clinical specimen; lane 4, negative control (distilled water); lane 5, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). (B) Results using hisJ primers. Lanes 1 and 9 contain molecular weight standards (100-bp DNA ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lanes 3 to 6, DNA extracted from culture-negative clinical specimens; lane 7, negative control (distilled water); lane 8, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). Arrows indicate amplicons produced in each reaction.
    Dna Sequences, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 94/100, based on 2081 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher dna sequencer
    <t>RT-PCR</t> analysis of ovarian tissues and OVCAR-3 cells. ( A ) RT-PCR analysis of ovarian serous carcinoma. Primers specific for the four known human SAA genes were used, and PCR fragments were analyzed on a 2% agarose gel. Markers of a <t>DNA</t> ladder (100-bp
    Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 8489 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Bioedit Company dna sequence
    Construction and analysis of <t>sreA</t> strains. A : Construction of pTFCM-R-L and pTFCM- neo - sreA plasmids. B: PCR analysis of Δ sreA , and CO sreA transformants using primers sreA -F and sreA -R ( Table 1 ). C: Southern blot analysis of P . digitatum wild-type PdHS-F6 and Δ sreA , CO sreA strains using a probe specific to the 5’ region of Pdsre . 30μg genomic <t>DNA</t> was digested with Hind III and detected using a probe specific to the 5’ region of sreA gene.
    Dna Sequence, supplied by Bioedit Company, used in various techniques. Bioz Stars score: 93/100, based on 107 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher dna sequence analysis
    Detection of the mRNA expression levels of IgG and related enzymes, including RAG1, RAG2 and AID in podocytes. (A) Transcripts of Igγ, γ1, γ3, γ4, κ and λ C regions were detected by <t>RT-PCR.</t> CD19 was detected as a marker of B lymphocytes. The mRNA of Igγ and κ V regions was amplified by nested RT-PCR. (B) mRNA expression levels of AID, RAG1 and RAG2. PBMCs were used as a positive control; water instead of cDNA was used as a blank control; R (cDNA template replaced by DNase-treated RNA) was used as negative control. (C) <t>DNA</t> sequencing results. The sequences of the PCR products were aligned with the mRNA sequences available in the NCBI database. HPC, human podocytes; AID, activation-induced cytidine deaminase; C, constant; Ig, immunoglobulin; Igγ, IgG heavy chain; PBMCs, peripheral blood mononuclear cells; RAG. recombination activating gene; RT-PCR, reverse transcription-polymerase chain reaction; V, variable.
    Dna Sequence Analysis, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1685 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi prism 377 dna sequencer
    Detection of the mRNA expression levels of IgG and related enzymes, including RAG1, RAG2 and AID in podocytes. (A) Transcripts of Igγ, γ1, γ3, γ4, κ and λ C regions were detected by <t>RT-PCR.</t> CD19 was detected as a marker of B lymphocytes. The mRNA of Igγ and κ V regions was amplified by nested RT-PCR. (B) mRNA expression levels of AID, RAG1 and RAG2. PBMCs were used as a positive control; water instead of cDNA was used as a blank control; R (cDNA template replaced by DNase-treated RNA) was used as negative control. (C) <t>DNA</t> sequencing results. The sequences of the PCR products were aligned with the mRNA sequences available in the NCBI database. HPC, human podocytes; AID, activation-induced cytidine deaminase; C, constant; Ig, immunoglobulin; Igγ, IgG heavy chain; PBMCs, peripheral blood mononuclear cells; RAG. recombination activating gene; RT-PCR, reverse transcription-polymerase chain reaction; V, variable.
    Abi Prism 377 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 3170 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa smart seq v4 ultra low input rna kit
    Comparison of FL-cDNA-Seq and Illumina <t>RNA-Seq.</t> (A) The gene expression of FL-cDNA-Seq of LC2/ad (R9.4) was compared with that of TruSeq RNA (left) and <t>SMART-Seq</t> (right). Pearson correlation coefficients are shown on the graph. (B) Influence of sequencing depth on the estimation of gene expression level and gene detection. Reads for each method were randomly sampled in triplicate. The average of the Pearson correlation coefficients between TruSeq RNA and randomly sampled data for FL-cDNA-Seq and SMART-Seq is shown (left). The average number of genes with an expression level of more than 1 tpm or ppm is shown (right). (C) Comparison to qRT-qPCR. Forty-four genes detected by all methods were analyzed. The gene expression of these genes was normalized to GAPDH. Pearson correlation coefficients are shown on the graph. qRT-qPCR data of LC2/ad was obtained as in our previous study. 16
    Smart Seq V4 Ultra Low Input Rna Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 1758 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Illumina Inc dna sequences
    Contents of biopolymers of the extracellular polymeric substances from <t>biofilm</t> samples harvested at T5. The <t>DNA</t> content was
    Dna Sequences, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 92/100, based on 483 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    GenScript dna sequences
    Large chromosomal deletions using two <t>TALENs.</t> (A) Schematic representation of the structure of partial BmBlos2 gene as described in Figure 3 . Red arrows at the top indicate primers used for PCR amplification. Blue lines at the bottom show the expected results of PCR amplification with (556 bp) or without (1351 bp) the large chromosomal deletion. (B) Gel analysis of PCR amplifications. WT represents the wild-type silkworm. Be-21 and Be-22 represent two G1 silkworm broods from G0 silkworms co-injected with B2 and B3, respectively. The numbers on the left represent the sizes of the <t>DNA</t> ladder (DL2000 plus). The blue arrows on the right indicate the two expected PCR products. (C) Sequences of the PCR products. The wild-type sequence is shown at the top with the full B2 site in red and the B3 site in blue. The TALEN recognition sites are underlined, and deletions are indicated by dashed lines.
    Dna Sequences, supplied by GenScript, used in various techniques. Bioz Stars score: 92/100, based on 590 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher abi 3730 dna sequencer
    Large chromosomal deletions using two <t>TALENs.</t> (A) Schematic representation of the structure of partial BmBlos2 gene as described in Figure 3 . Red arrows at the top indicate primers used for PCR amplification. Blue lines at the bottom show the expected results of PCR amplification with (556 bp) or without (1351 bp) the large chromosomal deletion. (B) Gel analysis of PCR amplifications. WT represents the wild-type silkworm. Be-21 and Be-22 represent two G1 silkworm broods from G0 silkworms co-injected with B2 and B3, respectively. The numbers on the left represent the sizes of the <t>DNA</t> ladder (DL2000 plus). The blue arrows on the right indicate the two expected PCR products. (C) Sequences of the PCR products. The wild-type sequence is shown at the top with the full B2 site in red and the B3 site in blue. The TALEN recognition sites are underlined, and deletions are indicated by dashed lines.
    Abi 3730 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2098 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc rna sequencing rna seq
    Changes in HESB expression in the brainstem of injured animals after drug treatments. (A) Changes in the expression of HESB in the brainstem of injured animals after <t>GABA</t> or baclofen treatments <t>(RNA-Seq).</t> HESB gene expression is represented as read counts normalized by DESeq’s median of ratios, and ** represents that the differences between the two groups are significant with false discovery rate corrected p -values between 0.01 and 0.001. Note that results are not comparable between the 2 RNA-Seq experiments. (B) Change in the expression of HESB in the brainstem of 1 wpl animals after the PF-3804014 treatment (qPCR). HESB expression is represented as log2 fold change in comparison to the mean of the controls, and * represents that the differences between the two groups are significant with p -value between 0.05 and 0.01. For both A and B, boxplots show the median, 25th and 75th percentiles, red dots represent the mean and whiskers extend to the most extreme data point which is no more than 1.5 times the length of the box away from the box.
    Rna Sequencing Rna Seq, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 99/100, based on 1269 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Genewiz dna sequence analysis
    The <t>bbb22</t> gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. <t>DNA</t> was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.
    Dna Sequence Analysis, supplied by Genewiz, used in various techniques. Bioz Stars score: 93/100, based on 311 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Eurofins dna sequences
    Telomere <t>DNA</t> G-quadruplex unfolding by arginine to alanine mutants of <t>UP1+RGG</t> monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.
    Dna Sequences, supplied by Eurofins, used in various techniques. Bioz Stars score: 92/100, based on 527 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc illumina dna sequencing
    Relationship between the <t>Illumina</t> <t>DNA-sequencing</t> read depth and the copy number inferred by ddPCR. (A) The Y-axis represents copy numbers per μl inferred by ddPCR. Black circles (gray background) and open circles (black background) indicate three-copy genes and single-copy genes, respectively. All points and error bars represent averages of four replicates and 95% CIs. (B) Each dot represents an A. halleri gene. The X-axis represents the Illumina DNA-sequencing read depth, which is the number of reads per 1 Kbp per 1 million reads. The Y-axis represents copy numbers per μl, which were inferred by ddPCR. The regression line was calculated with the simple formula Y = αX; α was inferred by the least squares method.
    Illumina Dna Sequencing, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 91/100, based on 950 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Thermo Fisher abi prism 3100 dna sequencer
    The <t>DNA</t> sequencing of rs840088. Direct sequencing of a subgroup of samples with the same primers for PCR-RFLP was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an <t>ABI</t> <t>PRISM</t> 3100 DNA sequencer (Applied Biosystems, Foster City, USA). (A) GG genotype. (B) GA genotype. (C) AA genotype. 82×48 mm (300×300 DPI).
    Abi Prism 3100 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 91/100, based on 1382 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Thermo Fisher abi 377 dna sequencer
    The <t>DNA</t> sequencing of rs840088. Direct sequencing of a subgroup of samples with the same primers for PCR-RFLP was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an <t>ABI</t> <t>PRISM</t> 3100 DNA sequencer (Applied Biosystems, Foster City, USA). (A) GG genotype. (B) GA genotype. (C) AA genotype. 82×48 mm (300×300 DPI).
    Abi 377 Dna Sequencer, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 1253 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Eurofins dna sequencing
    The <t>DNA</t> sequencing of rs840088. Direct sequencing of a subgroup of samples with the same primers for PCR-RFLP was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an <t>ABI</t> <t>PRISM</t> 3100 DNA sequencer (Applied Biosystems, Foster City, USA). (A) GG genotype. (B) GA genotype. (C) AA genotype. 82×48 mm (300×300 DPI).
    Dna Sequencing, supplied by Eurofins, used in various techniques. Bioz Stars score: 92/100, based on 4234 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Generation of a bb0028 mutant. A . An IPTG-regulatable bb0028 mutant was generated by inserting a streptomycin resistance cassette followed by the flacp promoter immediately upstream of bb0028 . Red arrows indicate primer-binding areas used to confirm insertion of the streptomycin cassette and flacp promoter into the borrelial genome. B . PCR amplification resulted in a 1.2 kb amplicon in the wildtype strain (WT), as expected, and a 3.0 kb amplicon in the BB0028 mutant (flacp::0028) indicating the streptomycin cassette and flacp promoter were inserted in the mutant. A PCR reaction with no DNA template (Neg) was included as a negative control. DNA sizes, in kilobase pairs, are indicated at left. C . Whole-cell lysate from the BB0028 mutant (flacp::0028) strain propagated with (+) or without (−) 1 mM IPTG, as well as from the parental, wildtype strain (WT), were subjected to immunoblot analysis using BB0028 specific antibodies (top panel) or FlaB antibodies (bottom panel). FlaB reactivity was used to ensure all lanes were loaded equally.

    Journal: BMC Microbiology

    Article Title: Characterization of the β-barrel assembly machine accessory lipoproteins from Borrelia burgdorferi

    doi: 10.1186/s12866-015-0411-y

    Figure Lengend Snippet: Generation of a bb0028 mutant. A . An IPTG-regulatable bb0028 mutant was generated by inserting a streptomycin resistance cassette followed by the flacp promoter immediately upstream of bb0028 . Red arrows indicate primer-binding areas used to confirm insertion of the streptomycin cassette and flacp promoter into the borrelial genome. B . PCR amplification resulted in a 1.2 kb amplicon in the wildtype strain (WT), as expected, and a 3.0 kb amplicon in the BB0028 mutant (flacp::0028) indicating the streptomycin cassette and flacp promoter were inserted in the mutant. A PCR reaction with no DNA template (Neg) was included as a negative control. DNA sizes, in kilobase pairs, are indicated at left. C . Whole-cell lysate from the BB0028 mutant (flacp::0028) strain propagated with (+) or without (−) 1 mM IPTG, as well as from the parental, wildtype strain (WT), were subjected to immunoblot analysis using BB0028 specific antibodies (top panel) or FlaB antibodies (bottom panel). FlaB reactivity was used to ensure all lanes were loaded equally.

    Article Snippet: The DNA sequence encoding mature BB0028, lacking the N-terminal leader peptide, was amplified from B. burgdorferi B31 genomic DNA using primers bb0028 (NheI) F and bb0028 (XhoI) R. The bamA P1-4 + 14 and bb0028 amplicons were then cloned into NheI and XhoI restriction sites of the the pET23a cloning vector (EMD Millipore, Darmstadt, Germany), and the resulting constructs were electroporated into E. coli Overexpress™ C41(DE3) (Lucigen Corp, Middleton, WI).

    Techniques: Mutagenesis, Generated, Binding Assay, Polymerase Chain Reaction, Amplification, Negative Control

    Validation of gene expression changes estimated with CANEapp with quantitative real-time PCR. a RNA-seq analysis of hippocampi of Alzheimer’s disease patients and controls. Hippocampal tissue from 4 AD patients and 4 control individuals was used to extract total RNA and perform ribodepletion and strand-specific library preparation. Single-end RNA sequencing was performed on Illumina HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes measured with real-time PCR was compared with expression values generated by CANEapp. b RNA-seq of developing mouse cortex. Tissue from 4 embryonic day 17 and 3 adult mouse cortical samples was processed to extract polyA-selected RNA and generate paired-end unidirectional sequencing data with Illumina Genome Analyzer IIx. Gene expression estimates of 4 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. c Fold changes of gene expression for RNA-seq of liver of rats treated with two DNA-damage compounds. The data was produced by paired-end sequencing of polyA-selected RNA on Illumina HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. R 2 -coefficient of determination

    Journal: BMC Genomics

    Article Title: CANEapp: a user-friendly application for automated next generation transcriptomic data analysis

    doi: 10.1186/s12864-015-2346-y

    Figure Lengend Snippet: Validation of gene expression changes estimated with CANEapp with quantitative real-time PCR. a RNA-seq analysis of hippocampi of Alzheimer’s disease patients and controls. Hippocampal tissue from 4 AD patients and 4 control individuals was used to extract total RNA and perform ribodepletion and strand-specific library preparation. Single-end RNA sequencing was performed on Illumina HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes measured with real-time PCR was compared with expression values generated by CANEapp. b RNA-seq of developing mouse cortex. Tissue from 4 embryonic day 17 and 3 adult mouse cortical samples was processed to extract polyA-selected RNA and generate paired-end unidirectional sequencing data with Illumina Genome Analyzer IIx. Gene expression estimates of 4 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. c Fold changes of gene expression for RNA-seq of liver of rats treated with two DNA-damage compounds. The data was produced by paired-end sequencing of polyA-selected RNA on Illumina HiSeq 2000. Fold changes of expression for 2 downregulated and 4 upregulated genes were compared between CANEapp and real-time PCR. R 2 -coefficient of determination

    Article Snippet: RNA in human and rat experiments was sequenced on the Illumina HiSeq 2000 machine, whereas mouse RNA-seq data was produced by sequencing RNA on Illumina GA-IIx sequencer.

    Techniques: Expressing, Real-time Polymerase Chain Reaction, RNA Sequencing Assay, Generated, Sequencing, Produced

    Genome-wide evaluation of mRNA stability states of expressed genes in RA FLS. (a-c), Gene tracks showing sequencing reads from RNA sequencing mapped to CCL20 (a), JUN (b) and IRF1 (c) genes. The sequencing reads after TNF stimulation for 1 hour without (blue) or with Act D (orange) are shown. (d), Stacked bar graphs illustrating the mRNA stability states of genes expressed in unstimulated (Control) and TNF-stimulated FLS (1, 3, 24 and 72 hours of TNF stimulation). The mRNA stability status was calculated as the ratio of expression levels at the TNF+Act D condition divided to the expression levels at the TNF condition. This ratio ranges from 0 to 1 and classifies genes to a spectrum from very unstable to very stable transcripts. The expressed genes were classified into five groups with distinct stability states and the size of each group is represented as % of total number of expressed genes for each condition.

    Journal: PLoS ONE

    Article Title: Tumor Necrosis Factor dynamically regulates the mRNA stabilome in rheumatoid arthritis fibroblast-like synoviocytes

    doi: 10.1371/journal.pone.0179762

    Figure Lengend Snippet: Genome-wide evaluation of mRNA stability states of expressed genes in RA FLS. (a-c), Gene tracks showing sequencing reads from RNA sequencing mapped to CCL20 (a), JUN (b) and IRF1 (c) genes. The sequencing reads after TNF stimulation for 1 hour without (blue) or with Act D (orange) are shown. (d), Stacked bar graphs illustrating the mRNA stability states of genes expressed in unstimulated (Control) and TNF-stimulated FLS (1, 3, 24 and 72 hours of TNF stimulation). The mRNA stability status was calculated as the ratio of expression levels at the TNF+Act D condition divided to the expression levels at the TNF condition. This ratio ranges from 0 to 1 and classifies genes to a spectrum from very unstable to very stable transcripts. The expressed genes were classified into five groups with distinct stability states and the size of each group is represented as % of total number of expressed genes for each condition.

    Article Snippet: RNA-sequencing libraries were generated from two patient donor FLS using Tru-Seq kits (Illumina) for poly-A selected transcripts.

    Techniques: Genome Wide, Sequencing, RNA Sequencing Assay, Activated Clotting Time Assay, Expressing

    Scatterplots comparing the expression levels to the mRNA stability states of the expressed genes in RA FLS. Two biological replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1, 3, 24, or 72 hours. Subsequently, actinomycin D (Act D, 10μg/ml) was added for 3 hours to block active transcription and gene expression was measured by RNA sequencing. RPKM values were generated using CuffDiff2. The mRNA stability status was calculated genome-wide as the ratio of RPKM levels at the TNF+Act D condition divided to the RPKM levels at the TNF condition. This ratio ranges from 0 to 1 and classifies genes to a spectrum from very unstable to very stable transcripts. The genes expressed at 1 (a), 3 (b), 24 (c), and 72 (d) hours of TNF stimulation were plotted based on their expression levels and the mRNA stability states. Shades of blue represent the region of unstable genes, and shades of red represent the zone of stable genes.

    Journal: PLoS ONE

    Article Title: Tumor Necrosis Factor dynamically regulates the mRNA stabilome in rheumatoid arthritis fibroblast-like synoviocytes

    doi: 10.1371/journal.pone.0179762

    Figure Lengend Snippet: Scatterplots comparing the expression levels to the mRNA stability states of the expressed genes in RA FLS. Two biological replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1, 3, 24, or 72 hours. Subsequently, actinomycin D (Act D, 10μg/ml) was added for 3 hours to block active transcription and gene expression was measured by RNA sequencing. RPKM values were generated using CuffDiff2. The mRNA stability status was calculated genome-wide as the ratio of RPKM levels at the TNF+Act D condition divided to the RPKM levels at the TNF condition. This ratio ranges from 0 to 1 and classifies genes to a spectrum from very unstable to very stable transcripts. The genes expressed at 1 (a), 3 (b), 24 (c), and 72 (d) hours of TNF stimulation were plotted based on their expression levels and the mRNA stability states. Shades of blue represent the region of unstable genes, and shades of red represent the zone of stable genes.

    Article Snippet: RNA-sequencing libraries were generated from two patient donor FLS using Tru-Seq kits (Illumina) for poly-A selected transcripts.

    Techniques: Expressing, Derivative Assay, Activated Clotting Time Assay, Blocking Assay, RNA Sequencing Assay, Generated, Genome Wide

    Association of expression kinetics with mRNA stability states of TNF-inducible genes in RA FLS. For (a-b), two biological replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1-72h. Subsequently, Act D (10 μg/ml) was added for 3h and gene expression was measured by RNA sequencing. 386 genes were identified as highly induced (≥5-fold) by TNF at any time point and were clustered into 6 clusters with distinct kinetics of peak expression. (a), Heatmap illustrating the expression kinetics of the 6 clusters (red represents the maximum and blue the minimum expression level across the lane). (b), Stacked bar graphs illustrating the stability states of genes for Cluster 1, Clusters 2 3, Cluster 4, and Clusters 5 6. For (c-f), RA FLS were exposed to a single dose of TNF (10 ng/ml) for 1–72 hours. Primers specific for the eighth intronic region of MMP3 and for the first intronic region of CCL5 were designed to capture primary transcripts (PT) of MMP3 and CCL5 . qPCR was used to measure the levels of PT and total mRNA of MMP3 (c-d) and CCL5 (e-f). Cumulative results from six independent experiments are shown. Values were normalized relative to mRNA for GAPDH and are presented as mean ±SEM.

    Journal: PLoS ONE

    Article Title: Tumor Necrosis Factor dynamically regulates the mRNA stabilome in rheumatoid arthritis fibroblast-like synoviocytes

    doi: 10.1371/journal.pone.0179762

    Figure Lengend Snippet: Association of expression kinetics with mRNA stability states of TNF-inducible genes in RA FLS. For (a-b), two biological replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1-72h. Subsequently, Act D (10 μg/ml) was added for 3h and gene expression was measured by RNA sequencing. 386 genes were identified as highly induced (≥5-fold) by TNF at any time point and were clustered into 6 clusters with distinct kinetics of peak expression. (a), Heatmap illustrating the expression kinetics of the 6 clusters (red represents the maximum and blue the minimum expression level across the lane). (b), Stacked bar graphs illustrating the stability states of genes for Cluster 1, Clusters 2 3, Cluster 4, and Clusters 5 6. For (c-f), RA FLS were exposed to a single dose of TNF (10 ng/ml) for 1–72 hours. Primers specific for the eighth intronic region of MMP3 and for the first intronic region of CCL5 were designed to capture primary transcripts (PT) of MMP3 and CCL5 . qPCR was used to measure the levels of PT and total mRNA of MMP3 (c-d) and CCL5 (e-f). Cumulative results from six independent experiments are shown. Values were normalized relative to mRNA for GAPDH and are presented as mean ±SEM.

    Article Snippet: RNA-sequencing libraries were generated from two patient donor FLS using Tru-Seq kits (Illumina) for poly-A selected transcripts.

    Techniques: Expressing, Derivative Assay, Activated Clotting Time Assay, RNA Sequencing Assay, Real-time Polymerase Chain Reaction

    Genome-wide identification of transcripts stabilized by TNF in RA FLS. Two biologic replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1 or 72h. Subsequently, Act D was added for 3h and gene expression was measured by RNA sequencing. The degree of TNF-induced mRNA stabilization was calculated as the log 2 difference of TNF+Act D/TNF ratio between 1 and 72h of TNF stimulation and the adjusted p values of TNF-induced stabilization were calculated by RiboDiff. (a), Scatter-plot of the genes displaying TNF-induced mRNA stabilization comparing the degree of mRNA stabilization (y axis) to the adjusted p values of the stabilizing effect of TNF (x-axis). (b), The top 40 genes displaying the highest TNF-induced mRNA stabilization ranked by the degree of stabilization. (c), Enriched biological processes identified by GSEA/MSigDB pathway analysis of the top 10% of the genes (n = 593) displaying the highest degree of TNF-induced mRNA stabilization.

    Journal: PLoS ONE

    Article Title: Tumor Necrosis Factor dynamically regulates the mRNA stabilome in rheumatoid arthritis fibroblast-like synoviocytes

    doi: 10.1371/journal.pone.0179762

    Figure Lengend Snippet: Genome-wide identification of transcripts stabilized by TNF in RA FLS. Two biologic replicates of RA FLS (derived from two different RA patients) were exposed to a single dose of TNF (10 ng/ml) for 1 or 72h. Subsequently, Act D was added for 3h and gene expression was measured by RNA sequencing. The degree of TNF-induced mRNA stabilization was calculated as the log 2 difference of TNF+Act D/TNF ratio between 1 and 72h of TNF stimulation and the adjusted p values of TNF-induced stabilization were calculated by RiboDiff. (a), Scatter-plot of the genes displaying TNF-induced mRNA stabilization comparing the degree of mRNA stabilization (y axis) to the adjusted p values of the stabilizing effect of TNF (x-axis). (b), The top 40 genes displaying the highest TNF-induced mRNA stabilization ranked by the degree of stabilization. (c), Enriched biological processes identified by GSEA/MSigDB pathway analysis of the top 10% of the genes (n = 593) displaying the highest degree of TNF-induced mRNA stabilization.

    Article Snippet: RNA-sequencing libraries were generated from two patient donor FLS using Tru-Seq kits (Illumina) for poly-A selected transcripts.

    Techniques: Genome Wide, Derivative Assay, Activated Clotting Time Assay, Expressing, RNA Sequencing Assay

    TNF induces expression of mRNA-stabilizing pathways and mRNA stabilization is MAPK-dependent. (a), RNA sequencing was performed in 2 biological replicates (derived from two different RA patients) of TNF-stimulated RA FLS and Panther-Gene Ontology was used to evaluate their enrichment for the biological process “Regulation of RNA stability” (GO:0043487 or GO:0043488). F.E = fold enrichment and ns = not significant. (b-h), RA FLS were exposed to a single dose of TNF (10 ng/ml) for 72h and then Act D (10 μg/ml) was added for 20 mins to block active transcription. Subsequently, the cells were treated for 4h with SB202190 (p38 inhibitor) alone or in various combinations with U0126 (MEK inhibitor) and SP600125 (JNK inhibitor). qPCR was used to measure the mRNA levels of CCL5 (b), IL-6 (c), IL-8 (d), CXCL3 (e), CCL2 (f), PTGS2 (g), and CXCL1 (h). Cumulative results from 4 independent experiments are shown. Values were normalized relative to GAPDH mRNA and presented as mean ±SEM. The mRNA expression at the TNF+Act D condition was set to 100 and the mRNA expression at all the other conditions was calculated as % of the TNF+Act D condition. P values were calculated by one-way ANOVA and Tukey post-test analysis (* = p

    Journal: PLoS ONE

    Article Title: Tumor Necrosis Factor dynamically regulates the mRNA stabilome in rheumatoid arthritis fibroblast-like synoviocytes

    doi: 10.1371/journal.pone.0179762

    Figure Lengend Snippet: TNF induces expression of mRNA-stabilizing pathways and mRNA stabilization is MAPK-dependent. (a), RNA sequencing was performed in 2 biological replicates (derived from two different RA patients) of TNF-stimulated RA FLS and Panther-Gene Ontology was used to evaluate their enrichment for the biological process “Regulation of RNA stability” (GO:0043487 or GO:0043488). F.E = fold enrichment and ns = not significant. (b-h), RA FLS were exposed to a single dose of TNF (10 ng/ml) for 72h and then Act D (10 μg/ml) was added for 20 mins to block active transcription. Subsequently, the cells were treated for 4h with SB202190 (p38 inhibitor) alone or in various combinations with U0126 (MEK inhibitor) and SP600125 (JNK inhibitor). qPCR was used to measure the mRNA levels of CCL5 (b), IL-6 (c), IL-8 (d), CXCL3 (e), CCL2 (f), PTGS2 (g), and CXCL1 (h). Cumulative results from 4 independent experiments are shown. Values were normalized relative to GAPDH mRNA and presented as mean ±SEM. The mRNA expression at the TNF+Act D condition was set to 100 and the mRNA expression at all the other conditions was calculated as % of the TNF+Act D condition. P values were calculated by one-way ANOVA and Tukey post-test analysis (* = p

    Article Snippet: RNA-sequencing libraries were generated from two patient donor FLS using Tru-Seq kits (Illumina) for poly-A selected transcripts.

    Techniques: Expressing, RNA Sequencing Assay, Derivative Assay, Activated Clotting Time Assay, Blocking Assay, Real-time Polymerase Chain Reaction

    Samples description and experimental design for next-generation sequencing. A ) 13 different tissues and conditions used for RNA purification. RAC : White young roots; FTN : Young leaves and stems; FTB : Leaves infected with Botrytis cinerea LR18; FTS: Leaves from water stressed plants; NDB: Dormant axillary buds (vegetative meristem); DBO: Active axillary buds (vegetative meristem); IFL: Floral bud at floral meristem transition; IMO: Floral meristem and early floral organs (sepal, petal, stamens and carpels) development; BFL: closed flower; DET: Stamens at microsporogenesis and microgametogenesis stages; OFT: open flower; SEN: senescent flower; CYN: rose hip from pollination up to early pigmentation. B , Sequencing and assembly strategy. Illumina reads were assembled using edena and combined with the trimmed 454 reads using TGICL to generate the final clusters assembly.

    Journal: BMC Genomics

    Article Title: Transcriptome database resource and gene expression atlas for the rose

    doi: 10.1186/1471-2164-13-638

    Figure Lengend Snippet: Samples description and experimental design for next-generation sequencing. A ) 13 different tissues and conditions used for RNA purification. RAC : White young roots; FTN : Young leaves and stems; FTB : Leaves infected with Botrytis cinerea LR18; FTS: Leaves from water stressed plants; NDB: Dormant axillary buds (vegetative meristem); DBO: Active axillary buds (vegetative meristem); IFL: Floral bud at floral meristem transition; IMO: Floral meristem and early floral organs (sepal, petal, stamens and carpels) development; BFL: closed flower; DET: Stamens at microsporogenesis and microgametogenesis stages; OFT: open flower; SEN: senescent flower; CYN: rose hip from pollination up to early pigmentation. B , Sequencing and assembly strategy. Illumina reads were assembled using edena and combined with the trimmed 454 reads using TGICL to generate the final clusters assembly.

    Article Snippet: 454 and Illumina sequencing RNA samples were checked for their integrity on an Agilent 2100 Bioanalyzer (Waldbroon, Germany) according to the manufacturer’s instructions.

    Techniques: Next-Generation Sequencing, Purification, Infection, Sequencing

    Transcriptomic mapping of genes associated with luteoloside biosynthesis in Lonicera macranthoides . Proposed pathways for luteoloside biosynthesis in Lonicera macranthoides were illustrated by RNA-Seq analysis. Luteolin, the precursor of luteoloside, is biosynthesized from the general flavonoid precursor: naringenin. Circle 1(①) indicates that luteolin is biosynthesized directly from apigenin catalyzed by F3’H. Circle 2(②) indicates that luteolin is generated directly from eriodictyol catalyzed by FNS. Expression profile for each gene was shown in colored blocks and each blocks represented the expression changes (represented by Log2Ratio) in senescing leaves with respect to young leaves. Red colors/green colors correspond to up-/down-regulation of these genes and Log2Ratio ≥ 1 is considered statistically significant. Details were showed in Table 2 . Abbreviations: PAL, phenylalanine ammonia lyase; C4H, cinnamate 4-hydroxylase; 4CL, 4-hydroxycinnamoyl CoA ligase/4-coumarate-CoA ligase; CHS, chalcone synthase; CHI, chalcone isomerase; FNS, flavone synthase; F3H, flavonoid 3′-monooxygenase/flavonoid 3′-hydroxylase; UF7GT, flavone 7- O -β-glucosyltransferase.

    Journal: International Journal of Molecular Sciences

    Article Title: Transcriptome Analysis Reveals Molecular Signatures of Luteoloside Accumulation in Senescing Leaves of Lonicera macranthoides

    doi: 10.3390/ijms19041012

    Figure Lengend Snippet: Transcriptomic mapping of genes associated with luteoloside biosynthesis in Lonicera macranthoides . Proposed pathways for luteoloside biosynthesis in Lonicera macranthoides were illustrated by RNA-Seq analysis. Luteolin, the precursor of luteoloside, is biosynthesized from the general flavonoid precursor: naringenin. Circle 1(①) indicates that luteolin is biosynthesized directly from apigenin catalyzed by F3’H. Circle 2(②) indicates that luteolin is generated directly from eriodictyol catalyzed by FNS. Expression profile for each gene was shown in colored blocks and each blocks represented the expression changes (represented by Log2Ratio) in senescing leaves with respect to young leaves. Red colors/green colors correspond to up-/down-regulation of these genes and Log2Ratio ≥ 1 is considered statistically significant. Details were showed in Table 2 . Abbreviations: PAL, phenylalanine ammonia lyase; C4H, cinnamate 4-hydroxylase; 4CL, 4-hydroxycinnamoyl CoA ligase/4-coumarate-CoA ligase; CHS, chalcone synthase; CHI, chalcone isomerase; FNS, flavone synthase; F3H, flavonoid 3′-monooxygenase/flavonoid 3′-hydroxylase; UF7GT, flavone 7- O -β-glucosyltransferase.

    Article Snippet: RNA Extraction and Illumina Sequencing Total RNA from young and senescing leaves was isolated and purified using QIAGEN RNeasy Plant Mini kit and RNase-free DNase set (QIAGEN, Hilden, Germany) according to manufacturer’s instruction.

    Techniques: RNA Sequencing Assay, Generated, Expressing

    Expression patterns of selected unigenes related to luteoloside biosynthesis identified by RNA-Seq were validated by qRT-PCR. Expressions of unigenes located upstream of luteoloside metabolic pathway ( A ) Unigene109136 ( PAL ), ( B ) CL11118.Contig2 ( C4H ), ( C ) Unigene29692 ( 4CL ) and downstream of luteoloside metabolic pathway ( D ) CL11269.Contig3 ( CHI ), ( E ) CL19869.Contig1 ( CHS ), ( F ) Unigene2335 ( FNSII ), ( G ) CL11828.Contig1 and ( H ) CL11828.Contig2 ( F3’H ), ( I ) Unigene2918 and ( J ) Unigene97915 ( UFGT ) in young and senescing leaves were analyzed by qRT-PCR. Relative expression levels were determined based on the reference young leaves set to 1. Values are means ± SD of three biological repetitions ( n = 3). Two technical replicates for each biological replicate were performed in each qRT-PCR experiments.

    Journal: International Journal of Molecular Sciences

    Article Title: Transcriptome Analysis Reveals Molecular Signatures of Luteoloside Accumulation in Senescing Leaves of Lonicera macranthoides

    doi: 10.3390/ijms19041012

    Figure Lengend Snippet: Expression patterns of selected unigenes related to luteoloside biosynthesis identified by RNA-Seq were validated by qRT-PCR. Expressions of unigenes located upstream of luteoloside metabolic pathway ( A ) Unigene109136 ( PAL ), ( B ) CL11118.Contig2 ( C4H ), ( C ) Unigene29692 ( 4CL ) and downstream of luteoloside metabolic pathway ( D ) CL11269.Contig3 ( CHI ), ( E ) CL19869.Contig1 ( CHS ), ( F ) Unigene2335 ( FNSII ), ( G ) CL11828.Contig1 and ( H ) CL11828.Contig2 ( F3’H ), ( I ) Unigene2918 and ( J ) Unigene97915 ( UFGT ) in young and senescing leaves were analyzed by qRT-PCR. Relative expression levels were determined based on the reference young leaves set to 1. Values are means ± SD of three biological repetitions ( n = 3). Two technical replicates for each biological replicate were performed in each qRT-PCR experiments.

    Article Snippet: RNA Extraction and Illumina Sequencing Total RNA from young and senescing leaves was isolated and purified using QIAGEN RNeasy Plant Mini kit and RNase-free DNase set (QIAGEN, Hilden, Germany) according to manufacturer’s instruction.

    Techniques: Expressing, RNA Sequencing Assay, Quantitative RT-PCR

    Number of differentially expressed unigenes during leaf senescence identified by RNA-Seq data in Lonicera macranthoides . Differentially expressed unigenes (DEGs) between young and senescing leaves were illustrated by bar chart. Red and green bars represent the up-regulated and down-regulated unigenes in senescing leaves (SL) compared with those in young leaves (YL), respectively in Lonicera macranthoides .

    Journal: International Journal of Molecular Sciences

    Article Title: Transcriptome Analysis Reveals Molecular Signatures of Luteoloside Accumulation in Senescing Leaves of Lonicera macranthoides

    doi: 10.3390/ijms19041012

    Figure Lengend Snippet: Number of differentially expressed unigenes during leaf senescence identified by RNA-Seq data in Lonicera macranthoides . Differentially expressed unigenes (DEGs) between young and senescing leaves were illustrated by bar chart. Red and green bars represent the up-regulated and down-regulated unigenes in senescing leaves (SL) compared with those in young leaves (YL), respectively in Lonicera macranthoides .

    Article Snippet: RNA Extraction and Illumina Sequencing Total RNA from young and senescing leaves was isolated and purified using QIAGEN RNeasy Plant Mini kit and RNase-free DNase set (QIAGEN, Hilden, Germany) according to manufacturer’s instruction.

    Techniques: RNA Sequencing Assay

    Expression patterns of selected unigenes related to transcription factors identified by RNA-Seq were validated by qRT-PCR. Expressions of unigenes, homologs to AtMYB12 ( A ) Unigene36582 and ( B ) Unigene52460 , homologs to VvMYB75 ( C ) CL5870.Contig1 , homologs to VvMYB1R1 ( D ) CL15118.Contig2 , homologs to AtbHLH113 ( E ) Unigene33227 and ( F ) Unigene11884 , homologs to AtbHLH78 ( G ) CL2495.Contig7 , homologs to TTG1 ( H ) Unigene108470 and homologs to AtMYB5 ( I ) CL20138.Contig2 in young and senescing leaves were analyzed by qRT-PCR. Relative expression levels were determined based on the reference young leaves set to 1. Values are means ± SD of three biological repetitions ( n = 3). Two technical replicates for each biological replicate were performed in each qRT-PCR experiments.

    Journal: International Journal of Molecular Sciences

    Article Title: Transcriptome Analysis Reveals Molecular Signatures of Luteoloside Accumulation in Senescing Leaves of Lonicera macranthoides

    doi: 10.3390/ijms19041012

    Figure Lengend Snippet: Expression patterns of selected unigenes related to transcription factors identified by RNA-Seq were validated by qRT-PCR. Expressions of unigenes, homologs to AtMYB12 ( A ) Unigene36582 and ( B ) Unigene52460 , homologs to VvMYB75 ( C ) CL5870.Contig1 , homologs to VvMYB1R1 ( D ) CL15118.Contig2 , homologs to AtbHLH113 ( E ) Unigene33227 and ( F ) Unigene11884 , homologs to AtbHLH78 ( G ) CL2495.Contig7 , homologs to TTG1 ( H ) Unigene108470 and homologs to AtMYB5 ( I ) CL20138.Contig2 in young and senescing leaves were analyzed by qRT-PCR. Relative expression levels were determined based on the reference young leaves set to 1. Values are means ± SD of three biological repetitions ( n = 3). Two technical replicates for each biological replicate were performed in each qRT-PCR experiments.

    Article Snippet: RNA Extraction and Illumina Sequencing Total RNA from young and senescing leaves was isolated and purified using QIAGEN RNeasy Plant Mini kit and RNase-free DNase set (QIAGEN, Hilden, Germany) according to manufacturer’s instruction.

    Techniques: Expressing, RNA Sequencing Assay, Quantitative RT-PCR

    Agarose gels of amplicons generated by PCR amplification of enriched fecal specimens using the invE-A (457-bp amplicon) and hisJ (496-bp amplicon) primer sets. (A) Results using the invE-A primers. Lanes 1 and 6 contain molecular weight standards (100-bp DNA ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lane 3, DNA extracted from culture-negative clinical specimen; lane 4, negative control (distilled water); lane 5, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). (B) Results using hisJ primers. Lanes 1 and 9 contain molecular weight standards (100-bp DNA ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lanes 3 to 6, DNA extracted from culture-negative clinical specimens; lane 7, negative control (distilled water); lane 8, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). Arrows indicate amplicons produced in each reaction.

    Journal: Journal of Clinical Microbiology

    Article Title: Identification of Two Phylogenetically Related Organisms from Feces by PCR for Detection of Salmonella spp.

    doi: 10.1128/JCM.40.4.1487-1492.2002

    Figure Lengend Snippet: Agarose gels of amplicons generated by PCR amplification of enriched fecal specimens using the invE-A (457-bp amplicon) and hisJ (496-bp amplicon) primer sets. (A) Results using the invE-A primers. Lanes 1 and 6 contain molecular weight standards (100-bp DNA ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lane 3, DNA extracted from culture-negative clinical specimen; lane 4, negative control (distilled water); lane 5, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). (B) Results using hisJ primers. Lanes 1 and 9 contain molecular weight standards (100-bp DNA ladder). DNA used for amplification was as follows: lane 2, DNA extracted from culture-positive clinical specimen; lanes 3 to 6, DNA extracted from culture-negative clinical specimens; lane 7, negative control (distilled water); lane 8, positive control (purified DNA from S. enterica serovar Infantis isolate 978-05817). Arrows indicate amplicons produced in each reaction.

    Article Snippet: To do this, the DNA sequences of the culture-negative and -positive amplicons and the negative and positive consensus sequences were submitted to the National Center for Biotechnology Information (NCBI) (National Library of Medicine, National Institutes of Health, Bethesda, Md.) for a nucleotide BLAST ( , ) search of the GenBank (NCBI), EMBL, DNA Databank of Japan (DDBJ), and Protein Data Bank (PDB) genetic sequence databases.

    Techniques: Generated, Polymerase Chain Reaction, Amplification, Molecular Weight, Negative Control, Positive Control, Purification, Produced

    RT-PCR analysis of ovarian tissues and OVCAR-3 cells. ( A ) RT-PCR analysis of ovarian serous carcinoma. Primers specific for the four known human SAA genes were used, and PCR fragments were analyzed on a 2% agarose gel. Markers of a DNA ladder (100-bp

    Journal: Journal of Histochemistry and Cytochemistry

    Article Title: Expression of Serum Amyloid A in Human Ovarian Epithelial Tumors: Implication for a Role in Ovarian Tumorigenesis

    doi: 10.1369/jhc.2010.956821

    Figure Lengend Snippet: RT-PCR analysis of ovarian tissues and OVCAR-3 cells. ( A ) RT-PCR analysis of ovarian serous carcinoma. Primers specific for the four known human SAA genes were used, and PCR fragments were analyzed on a 2% agarose gel. Markers of a DNA ladder (100-bp

    Article Snippet: The identity of the PCR products was confirmed by their sequencing in an automated DNA sequencer (Applied Biosystems 3730 DNA Analyzer; Hy Laboratories; Rehovot, Israel).

    Techniques: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Agarose Gel Electrophoresis

    Sequence of the FGF-2 mRNA leader. (A) The sequence of the FGF-2 cDNA 5′ region was obtained by using a DNA analyzer (see Materials and Methods) and compared to the two published sequences. Under the schema of the FGF-2 cDNA indicating the two Hga ). Line 3 is the sequence obtained in this study for both DNAs with the DNA analyzer. The new Hga I site is underlined (position 153). (B) The pFC1 plasmid with the 5′ region of the FGF-2 cDNA, its homolog with the corresponding genomic sequence (kindly provided by R. Florkiewicz), and a pFC1-derived plasmid lacking the Hga ]) were Bss HII digested. The 291-nt-long resulting fragments were dephosphorylated, kinase treated in the presence of [ 32 P]ATP, and then digested with enzyme Hga I. Each step was followed by a G50 column purification. The restriction fragments were fractionated on a 15% polyacrylamide–Tris-borate-EDTA gel, which was dried and autoradiographed. Sizes of the expected fragments are (i) 121, 85, 51, and 34 nt in the presence of the new Hga I site and (ii) 121 nt plus a doublet at 85 nt in its absence. The DNA origin (cDNA or genomic) is indicated at the top; control corresponds to the plasmid lacking the Hga I site at position 153. (C) Representation of the new reading frame given by the DNA sequence shown in A (line 3). The sequence between nt 80 and 163 is shown. The two potential initiation codons are shown in boldface; the Hga I site (positions 153 to 157) is underlined.

    Journal: Molecular and Cellular Biology

    Article Title: A New 34-Kilodalton Isoform of Human Fibroblast Growth Factor 2 Is Cap Dependently Synthesized by Using a Non-AUG Start Codon and Behaves as a Survival Factor

    doi:

    Figure Lengend Snippet: Sequence of the FGF-2 mRNA leader. (A) The sequence of the FGF-2 cDNA 5′ region was obtained by using a DNA analyzer (see Materials and Methods) and compared to the two published sequences. Under the schema of the FGF-2 cDNA indicating the two Hga ). Line 3 is the sequence obtained in this study for both DNAs with the DNA analyzer. The new Hga I site is underlined (position 153). (B) The pFC1 plasmid with the 5′ region of the FGF-2 cDNA, its homolog with the corresponding genomic sequence (kindly provided by R. Florkiewicz), and a pFC1-derived plasmid lacking the Hga ]) were Bss HII digested. The 291-nt-long resulting fragments were dephosphorylated, kinase treated in the presence of [ 32 P]ATP, and then digested with enzyme Hga I. Each step was followed by a G50 column purification. The restriction fragments were fractionated on a 15% polyacrylamide–Tris-borate-EDTA gel, which was dried and autoradiographed. Sizes of the expected fragments are (i) 121, 85, 51, and 34 nt in the presence of the new Hga I site and (ii) 121 nt plus a doublet at 85 nt in its absence. The DNA origin (cDNA or genomic) is indicated at the top; control corresponds to the plasmid lacking the Hga I site at position 153. (C) Representation of the new reading frame given by the DNA sequence shown in A (line 3). The sequence between nt 80 and 163 is shown. The two potential initiation codons are shown in boldface; the Hga I site (positions 153 to 157) is underlined.

    Article Snippet: By sequencing this region of the FGF-2 cDNA with an automatic DNA sequencer (Applied Biosystems), we obtained the DNA sequence 153-GACGCGGT downstream from position 153, which still differed from the published sequences of cDNA and genomic DNA, i.e., 153-GACGGCT and 153-GACGGT, respectively (Fig. A, line 3).

    Techniques: Sequencing, Plasmid Preparation, Derivative Assay, Purification

    Analysis of the ability of the FGF-2 mRNA 5′ UTR to promote translation initiation of the 34-kDa FGF-2. Expression of the 34-kDa FGF-2 was analyzed for three DNA constructs either by in vitro transcription and translation in RRL or by COS-7 cell transfection followed by Western immunoblotting (see Materials and Methods). (A) Schema of the mRNAs expressed from the different constructs (see Materials and Methods). WT5′-FGF corresponds to the FGF-2 mRNA with a complete 5′ leader; Δ1-257 corresponds to a deletion of nt 1 to 257 in the FGF-2 mRNA 5′ end; WT5′CAT corresponds to a fusion of FGF-2 cDNA nt 1 to 312 with the CAT sequence (devoid of an AUG start codon). (B) Analysis of mRNA expression either by in vitro translation (lanes 1 and 2) or by Western immunoblotting (lanes 3 to 6). Names of the antibodies (αFGF-2, αCAT, and αN23) are indicated at the bottom. Migration of size standards is indicated on the right; migration of FGF-2 isoforms is indicated on the left.

    Journal: Molecular and Cellular Biology

    Article Title: A New 34-Kilodalton Isoform of Human Fibroblast Growth Factor 2 Is Cap Dependently Synthesized by Using a Non-AUG Start Codon and Behaves as a Survival Factor

    doi:

    Figure Lengend Snippet: Analysis of the ability of the FGF-2 mRNA 5′ UTR to promote translation initiation of the 34-kDa FGF-2. Expression of the 34-kDa FGF-2 was analyzed for three DNA constructs either by in vitro transcription and translation in RRL or by COS-7 cell transfection followed by Western immunoblotting (see Materials and Methods). (A) Schema of the mRNAs expressed from the different constructs (see Materials and Methods). WT5′-FGF corresponds to the FGF-2 mRNA with a complete 5′ leader; Δ1-257 corresponds to a deletion of nt 1 to 257 in the FGF-2 mRNA 5′ end; WT5′CAT corresponds to a fusion of FGF-2 cDNA nt 1 to 312 with the CAT sequence (devoid of an AUG start codon). (B) Analysis of mRNA expression either by in vitro translation (lanes 1 and 2) or by Western immunoblotting (lanes 3 to 6). Names of the antibodies (αFGF-2, αCAT, and αN23) are indicated at the bottom. Migration of size standards is indicated on the right; migration of FGF-2 isoforms is indicated on the left.

    Article Snippet: By sequencing this region of the FGF-2 cDNA with an automatic DNA sequencer (Applied Biosystems), we obtained the DNA sequence 153-GACGCGGT downstream from position 153, which still differed from the published sequences of cDNA and genomic DNA, i.e., 153-GACGGCT and 153-GACGGT, respectively (Fig. A, line 3).

    Techniques: Expressing, Construct, In Vitro, Transfection, Western Blot, Sequencing, Migration

    Construction and analysis of sreA strains. A : Construction of pTFCM-R-L and pTFCM- neo - sreA plasmids. B: PCR analysis of Δ sreA , and CO sreA transformants using primers sreA -F and sreA -R ( Table 1 ). C: Southern blot analysis of P . digitatum wild-type PdHS-F6 and Δ sreA , CO sreA strains using a probe specific to the 5’ region of Pdsre . 30μg genomic DNA was digested with Hind III and detected using a probe specific to the 5’ region of sreA gene.

    Journal: PLoS ONE

    Article Title: A Novel Sterol Regulatory Element-Binding Protein Gene (sreA) Identified in Penicilliumdigitatum Is Required for Prochloraz Resistance, Full Virulence and erg11 (cyp51) Regulation

    doi: 10.1371/journal.pone.0117115

    Figure Lengend Snippet: Construction and analysis of sreA strains. A : Construction of pTFCM-R-L and pTFCM- neo - sreA plasmids. B: PCR analysis of Δ sreA , and CO sreA transformants using primers sreA -F and sreA -R ( Table 1 ). C: Southern blot analysis of P . digitatum wild-type PdHS-F6 and Δ sreA , CO sreA strains using a probe specific to the 5’ region of Pdsre . 30μg genomic DNA was digested with Hind III and detected using a probe specific to the 5’ region of sreA gene.

    Article Snippet: DNA and protein analysis The DNA sequence of sreA was analyzed using NCBI BLAST and BioEdit software.

    Techniques: Polymerase Chain Reaction, Southern Blot

    Detection of the mRNA expression levels of IgG and related enzymes, including RAG1, RAG2 and AID in podocytes. (A) Transcripts of Igγ, γ1, γ3, γ4, κ and λ C regions were detected by RT-PCR. CD19 was detected as a marker of B lymphocytes. The mRNA of Igγ and κ V regions was amplified by nested RT-PCR. (B) mRNA expression levels of AID, RAG1 and RAG2. PBMCs were used as a positive control; water instead of cDNA was used as a blank control; R (cDNA template replaced by DNase-treated RNA) was used as negative control. (C) DNA sequencing results. The sequences of the PCR products were aligned with the mRNA sequences available in the NCBI database. HPC, human podocytes; AID, activation-induced cytidine deaminase; C, constant; Ig, immunoglobulin; Igγ, IgG heavy chain; PBMCs, peripheral blood mononuclear cells; RAG. recombination activating gene; RT-PCR, reverse transcription-polymerase chain reaction; V, variable.

    Journal: International Journal of Molecular Medicine

    Article Title: Expression of immunoglobulin G in human podocytes, and its role in cell viability and adhesion

    doi: 10.3892/ijmm.2018.3525

    Figure Lengend Snippet: Detection of the mRNA expression levels of IgG and related enzymes, including RAG1, RAG2 and AID in podocytes. (A) Transcripts of Igγ, γ1, γ3, γ4, κ and λ C regions were detected by RT-PCR. CD19 was detected as a marker of B lymphocytes. The mRNA of Igγ and κ V regions was amplified by nested RT-PCR. (B) mRNA expression levels of AID, RAG1 and RAG2. PBMCs were used as a positive control; water instead of cDNA was used as a blank control; R (cDNA template replaced by DNase-treated RNA) was used as negative control. (C) DNA sequencing results. The sequences of the PCR products were aligned with the mRNA sequences available in the NCBI database. HPC, human podocytes; AID, activation-induced cytidine deaminase; C, constant; Ig, immunoglobulin; Igγ, IgG heavy chain; PBMCs, peripheral blood mononuclear cells; RAG. recombination activating gene; RT-PCR, reverse transcription-polymerase chain reaction; V, variable.

    Article Snippet: The reliability of PCR products was analyzed by DNA sequence analysis, which was performed by Invitrogen Trading (Shanghai) Co., Ltd. (Shanghai, China).

    Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Marker, Amplification, Positive Control, Negative Control, DNA Sequencing, Polymerase Chain Reaction, Activation Assay

    Comparison of FL-cDNA-Seq and Illumina RNA-Seq. (A) The gene expression of FL-cDNA-Seq of LC2/ad (R9.4) was compared with that of TruSeq RNA (left) and SMART-Seq (right). Pearson correlation coefficients are shown on the graph. (B) Influence of sequencing depth on the estimation of gene expression level and gene detection. Reads for each method were randomly sampled in triplicate. The average of the Pearson correlation coefficients between TruSeq RNA and randomly sampled data for FL-cDNA-Seq and SMART-Seq is shown (left). The average number of genes with an expression level of more than 1 tpm or ppm is shown (right). (C) Comparison to qRT-qPCR. Forty-four genes detected by all methods were analyzed. The gene expression of these genes was normalized to GAPDH. Pearson correlation coefficients are shown on the graph. qRT-qPCR data of LC2/ad was obtained as in our previous study. 16

    Journal: DNA Research: An International Journal for Rapid Publication of Reports on Genes and Genomes

    Article Title: Evaluation and application of RNA-Seq by MinION

    doi: 10.1093/dnares/dsy038

    Figure Lengend Snippet: Comparison of FL-cDNA-Seq and Illumina RNA-Seq. (A) The gene expression of FL-cDNA-Seq of LC2/ad (R9.4) was compared with that of TruSeq RNA (left) and SMART-Seq (right). Pearson correlation coefficients are shown on the graph. (B) Influence of sequencing depth on the estimation of gene expression level and gene detection. Reads for each method were randomly sampled in triplicate. The average of the Pearson correlation coefficients between TruSeq RNA and randomly sampled data for FL-cDNA-Seq and SMART-Seq is shown (left). The average number of genes with an expression level of more than 1 tpm or ppm is shown (right). (C) Comparison to qRT-qPCR. Forty-four genes detected by all methods were analyzed. The gene expression of these genes was normalized to GAPDH. Pearson correlation coefficients are shown on the graph. qRT-qPCR data of LC2/ad was obtained as in our previous study. 16

    Article Snippet: Optimization of the cDNA-Seq procedure Using the SMART-Seq v4 Ultra Low Input RNA Kit from Takara Bio, which is based on the Smart-Seq2 method, we synthesized full-length cDNA (FL-cDNA) from 50 ng total RNA isolated from seven lung adenocarcinoma-derived cell lines: PC-7, PC-9, H1975, H2228, VMRC-LCD, LC2/ad, and A549 ( ).

    Techniques: RNA Sequencing Assay, Expressing, Sequencing, Real-time Polymerase Chain Reaction

    Contents of biopolymers of the extracellular polymeric substances from biofilm samples harvested at T5. The DNA content was

    Journal: Applied and Environmental Microbiology

    Article Title: Metagenome Survey of a Multispecies and Alga-Associated Biofilm Revealed Key Elements of Bacterial-Algal Interactions in Photobioreactors

    doi: 10.1128/AEM.01641-13

    Figure Lengend Snippet: Contents of biopolymers of the extracellular polymeric substances from biofilm samples harvested at T5. The DNA content was

    Article Snippet: To further verify these data, we analyzed the DNA sequences of the mature PBR biofilm obtained by pyrosequencing and Illumina-based sequencing for the presence of hypervariable regions of rRNA gene fragments.

    Techniques:

    Large chromosomal deletions using two TALENs. (A) Schematic representation of the structure of partial BmBlos2 gene as described in Figure 3 . Red arrows at the top indicate primers used for PCR amplification. Blue lines at the bottom show the expected results of PCR amplification with (556 bp) or without (1351 bp) the large chromosomal deletion. (B) Gel analysis of PCR amplifications. WT represents the wild-type silkworm. Be-21 and Be-22 represent two G1 silkworm broods from G0 silkworms co-injected with B2 and B3, respectively. The numbers on the left represent the sizes of the DNA ladder (DL2000 plus). The blue arrows on the right indicate the two expected PCR products. (C) Sequences of the PCR products. The wild-type sequence is shown at the top with the full B2 site in red and the B3 site in blue. The TALEN recognition sites are underlined, and deletions are indicated by dashed lines.

    Journal: PLoS ONE

    Article Title: Highly Efficient and Specific Genome Editing in Silkworm Using Custom TALENs

    doi: 10.1371/journal.pone.0045035

    Figure Lengend Snippet: Large chromosomal deletions using two TALENs. (A) Schematic representation of the structure of partial BmBlos2 gene as described in Figure 3 . Red arrows at the top indicate primers used for PCR amplification. Blue lines at the bottom show the expected results of PCR amplification with (556 bp) or without (1351 bp) the large chromosomal deletion. (B) Gel analysis of PCR amplifications. WT represents the wild-type silkworm. Be-21 and Be-22 represent two G1 silkworm broods from G0 silkworms co-injected with B2 and B3, respectively. The numbers on the left represent the sizes of the DNA ladder (DL2000 plus). The blue arrows on the right indicate the two expected PCR products. (C) Sequences of the PCR products. The wild-type sequence is shown at the top with the full B2 site in red and the B3 site in blue. The TALEN recognition sites are underlined, and deletions are indicated by dashed lines.

    Article Snippet: The designed amino acid sequences of TALENs (designated as B2L, B2R, B3L, and B3R) were converted to DNA sequences that correspond with the codon usage in silkworm and then synthesized using a commercial service (GenScript).

    Techniques: TALENs, Polymerase Chain Reaction, Amplification, Injection, Sequencing

    Design and construction of TALENs that target the BmBlos2 gene. (A) Schematic representation of the BmBlos2 gene structure depicting the introns (broken lines), exons (colored boxes) and TALEN target sequences (sequences at the bottom). The numbers indicate the exact lengths of the introns and exons. ATG and TAA indicate the translation start and stop sites, respectively. The underlined sequence represents the recognition site of the corresponding TALEN monomer. (B) Schematic representation of TALEN binding to its target DNA. A triple flag tag and a nuclear localization signal (NLS) (grey boxes) are fused to the N-terminal of each TALEN monomer. The Fok I domain (red boxes) is linked to the C-terminal of each TALEN monomer through a flexible linker. The TAL effector domain is composed of an N-terminal (blue box labelled NT), a C-terminal (blue box labelled CT), and a tandem array of repeats (an array of colored boxes between NT and CT). The TALEN binding site is composed of a left binding site (underlined), right binding site (underlined) and a spacer (NNN…). (C) TALEN target sequences and the corresponding RVDs within each repeat. The number at the top of the left panel indicates the position of the repeat. The letters at the top of each TALEN monomer represent the target sequence and the letters below represent the RVDs of the corresponding repeat. The right panel represents the RVD usage in the design. The yellow coloured RVDs are different from the commonly used RVDs.

    Journal: PLoS ONE

    Article Title: Highly Efficient and Specific Genome Editing in Silkworm Using Custom TALENs

    doi: 10.1371/journal.pone.0045035

    Figure Lengend Snippet: Design and construction of TALENs that target the BmBlos2 gene. (A) Schematic representation of the BmBlos2 gene structure depicting the introns (broken lines), exons (colored boxes) and TALEN target sequences (sequences at the bottom). The numbers indicate the exact lengths of the introns and exons. ATG and TAA indicate the translation start and stop sites, respectively. The underlined sequence represents the recognition site of the corresponding TALEN monomer. (B) Schematic representation of TALEN binding to its target DNA. A triple flag tag and a nuclear localization signal (NLS) (grey boxes) are fused to the N-terminal of each TALEN monomer. The Fok I domain (red boxes) is linked to the C-terminal of each TALEN monomer through a flexible linker. The TAL effector domain is composed of an N-terminal (blue box labelled NT), a C-terminal (blue box labelled CT), and a tandem array of repeats (an array of colored boxes between NT and CT). The TALEN binding site is composed of a left binding site (underlined), right binding site (underlined) and a spacer (NNN…). (C) TALEN target sequences and the corresponding RVDs within each repeat. The number at the top of the left panel indicates the position of the repeat. The letters at the top of each TALEN monomer represent the target sequence and the letters below represent the RVDs of the corresponding repeat. The right panel represents the RVD usage in the design. The yellow coloured RVDs are different from the commonly used RVDs.

    Article Snippet: The designed amino acid sequences of TALENs (designated as B2L, B2R, B3L, and B3R) were converted to DNA sequences that correspond with the codon usage in silkworm and then synthesized using a commercial service (GenScript).

    Techniques: TALENs, Sequencing, Binding Assay, FLAG-tag

    Changes in HESB expression in the brainstem of injured animals after drug treatments. (A) Changes in the expression of HESB in the brainstem of injured animals after GABA or baclofen treatments (RNA-Seq). HESB gene expression is represented as read counts normalized by DESeq’s median of ratios, and ** represents that the differences between the two groups are significant with false discovery rate corrected p -values between 0.01 and 0.001. Note that results are not comparable between the 2 RNA-Seq experiments. (B) Change in the expression of HESB in the brainstem of 1 wpl animals after the PF-3804014 treatment (qPCR). HESB expression is represented as log2 fold change in comparison to the mean of the controls, and * represents that the differences between the two groups are significant with p -value between 0.05 and 0.01. For both A and B, boxplots show the median, 25th and 75th percentiles, red dots represent the mean and whiskers extend to the most extreme data point which is no more than 1.5 times the length of the box away from the box.

    Journal: Frontiers in Cell and Developmental Biology

    Article Title: Inhibition of Gamma-Secretase Promotes Axon Regeneration After a Complete Spinal Cord Injury

    doi: 10.3389/fcell.2020.00173

    Figure Lengend Snippet: Changes in HESB expression in the brainstem of injured animals after drug treatments. (A) Changes in the expression of HESB in the brainstem of injured animals after GABA or baclofen treatments (RNA-Seq). HESB gene expression is represented as read counts normalized by DESeq’s median of ratios, and ** represents that the differences between the two groups are significant with false discovery rate corrected p -values between 0.01 and 0.001. Note that results are not comparable between the 2 RNA-Seq experiments. (B) Change in the expression of HESB in the brainstem of 1 wpl animals after the PF-3804014 treatment (qPCR). HESB expression is represented as log2 fold change in comparison to the mean of the controls, and * represents that the differences between the two groups are significant with p -value between 0.05 and 0.01. For both A and B, boxplots show the median, 25th and 75th percentiles, red dots represent the mean and whiskers extend to the most extreme data point which is no more than 1.5 times the length of the box away from the box.

    Article Snippet: Here, we repeated the GABA and baclofen treatments and carried out 2 independent Illumina RNA sequencing (RNA-Seq) studies in the brainstems of control non-treated animals and treated animals.

    Techniques: Expressing, RNA Sequencing Assay, Real-time Polymerase Chain Reaction

    The bbb22 gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. DNA was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.

    Journal: Infection and Immunity

    Article Title: Molecular Dissection of a Borrelia burgdorferiIn Vivo Essential Purine Transport System

    doi: 10.1128/IAI.02859-14

    Figure Lengend Snippet: The bbb22 gene alone is not sufficient to maintain wild-type levels of spirochete loads in infected mouse tissues. DNA was isolated from ear, heart, and joint tissues of C3H/HeN mice inoculated with 1 × 10 4 bbb22-23 + or 22dist p -bbb22 + spirochetes.

    Article Snippet: Plasmids pBSV2G 22dist p - bbb22 + and pBSV2G 22prox p - bbb22 + were confirmed by restriction digestion and DNA sequence analysis (Genewiz).

    Techniques: Infection, Isolation, Mouse Assay

    Telomere DNA G-quadruplex unfolding by arginine to alanine mutants of UP1+RGG monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by arginine to alanine mutants of UP1+RGG monitored using CD spectroscopy. ( A, B ) Unfolding of K + form of Tel22 G-quadruplex DNA by TriRGG (A) and AllRGG (B) mutants. The G-quadruplex DNA was titrated with increasing molar excess of proteins. The black arrows in the spectra indicate the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C ) The normalized ellipticity at 295 nm of Tel22 at the final titration step (at 1:6 molar ratio of DNA to protein) for UP1, AllRGG, TriRGG and UP1+RGG showing the relative foldedness of the G-quadruplex structure.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Spectroscopy, Protein Concentration, Titration

    Interaction of RGG-box with the single stranded and G-quadruplex DNA monitored through NMR spectroscopy. ( A ) 2D 15 N– 1 H HSQC spectrum of the free RGG-box (black) and in complex with Tel22ss at 1:6 protein to DNA molar ratio (red). No significant chemical shift perturbations were observed for this interaction. Single stranded Tel22ss is shown as a cartoon. ( B ) 2D 15 N– 1 H HSQC spectrum of the RGG-box (black) and in complex with Tel22 at 1:6 protein to DNA molar ratio (red). Specific chemical shift perturbations were observed for several residues (marked with green arrows). A representative cartoon of monomeric G-quadruplex form of Tel22 is shown (only one conformation in K + ion is shown). ( C ) A subset of residues of RGG-box that show specific chemical shift perturbation upon addition of Tel22 is shown. The RGG-box and Tel22 complex was in fast exchange (weak binding) as we observed continuous movement of resonance peaks upon addition of increasing amount of the Tel22 DNA. Three steps of titration at different protein to DNA ratios are shown: black at 1:0, green at 1: 1, and red at 1:6. ( D ) The titration curves showing chemical shift change plotted as a function of increasing DNA:protein ratio for 14 interacting residues of the RGG-box is shown.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Interaction of RGG-box with the single stranded and G-quadruplex DNA monitored through NMR spectroscopy. ( A ) 2D 15 N– 1 H HSQC spectrum of the free RGG-box (black) and in complex with Tel22ss at 1:6 protein to DNA molar ratio (red). No significant chemical shift perturbations were observed for this interaction. Single stranded Tel22ss is shown as a cartoon. ( B ) 2D 15 N– 1 H HSQC spectrum of the RGG-box (black) and in complex with Tel22 at 1:6 protein to DNA molar ratio (red). Specific chemical shift perturbations were observed for several residues (marked with green arrows). A representative cartoon of monomeric G-quadruplex form of Tel22 is shown (only one conformation in K + ion is shown). ( C ) A subset of residues of RGG-box that show specific chemical shift perturbation upon addition of Tel22 is shown. The RGG-box and Tel22 complex was in fast exchange (weak binding) as we observed continuous movement of resonance peaks upon addition of increasing amount of the Tel22 DNA. Three steps of titration at different protein to DNA ratios are shown: black at 1:0, green at 1: 1, and red at 1:6. ( D ) The titration curves showing chemical shift change plotted as a function of increasing DNA:protein ratio for 14 interacting residues of the RGG-box is shown.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Nuclear Magnetic Resonance, Spectroscopy, Binding Assay, Titration

    Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using CD spectroscopy. ( A, B ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C, D ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1+RGG. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( E, F ) The ellipticity at 295 nm was normalized and plotted to show the relative foldedness of both K + and Na + forms of quadruplexes upon UP1 or UP1+RGG addition at each step of titration.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using CD spectroscopy. ( A, B ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( C, D ) Unfolding of K + and Na + forms of Tel22 G-quadruplex DNA by UP1+RGG. The G-quadruplex DNAs were titrated with increasing molar excess of proteins. The black arrow in the spectra indicates the gradual decrease in ellipticity at 295 nm with increasing protein concentration. ( E, F ) The ellipticity at 295 nm was normalized and plotted to show the relative foldedness of both K + and Na + forms of quadruplexes upon UP1 or UP1+RGG addition at each step of titration.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Spectroscopy, Protein Concentration, Titration

    Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using NMR and fluorescence spectroscopy. ( A ) 1D 1 H NMR spectra of Na + form of Tel22 showing gradual loss of imino proton peaks upon titration with increasing concentrations of UP1 (blue) and UP1+RGG (red). ( B ) Unfolding of the 5′-FAM and 3′-TAMRA labeled K + form of Tel22 DNA G-quadruplex (5′FAM-Tel22-TAMRA3′) by UP1 (blue) and UP1+RGG (red) monitored by observing the emission of FAM at 516 nM. 5′FAM-Tel22-TAMRA3′ DNA was mixed with 4 molar equivalents of UP1 or UP1+RGG and the emission spectrum was recorded over a time period. ( C ) Proposed model for RGG-box assisted recognition and unfolding of telomere DNA G-quadruplex unfolding by UP1+RGG.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Telomere DNA G-quadruplex unfolding by UP1+RGG and UP1 monitored using NMR and fluorescence spectroscopy. ( A ) 1D 1 H NMR spectra of Na + form of Tel22 showing gradual loss of imino proton peaks upon titration with increasing concentrations of UP1 (blue) and UP1+RGG (red). ( B ) Unfolding of the 5′-FAM and 3′-TAMRA labeled K + form of Tel22 DNA G-quadruplex (5′FAM-Tel22-TAMRA3′) by UP1 (blue) and UP1+RGG (red) monitored by observing the emission of FAM at 516 nM. 5′FAM-Tel22-TAMRA3′ DNA was mixed with 4 molar equivalents of UP1 or UP1+RGG and the emission spectrum was recorded over a time period. ( C ) Proposed model for RGG-box assisted recognition and unfolding of telomere DNA G-quadruplex unfolding by UP1+RGG.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques: Nuclear Magnetic Resonance, Fluorescence, Spectroscopy, Titration, Labeling

    Interaction of UP1+RGG and UP1 with the single stranded and G-quadruplex DNA monitored through ITC. Raw and fitted isotherms are shown and the equilibrium K d s obtained upon fitting of the raw data is mentioned in each panel. ( A, B ) Interaction of UP1 with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( C, D ) Interaction of UP1+RGG with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( E, F ) Interaction of UP1 with the Na + and K + forms of Tel22 G-quadruplex DNA respectively. ( G, H ) Interaction of UP1+RGG with the Na + and K + forms of Tel22 G-quadruplex DNA respectively.

    Journal: Nucleic Acids Research

    Article Title: RGG-box in hnRNPA1 specifically recognizes the telomere G-quadruplex DNA and enhances the G-quadruplex unfolding ability of UP1 domain

    doi: 10.1093/nar/gky854

    Figure Lengend Snippet: Interaction of UP1+RGG and UP1 with the single stranded and G-quadruplex DNA monitored through ITC. Raw and fitted isotherms are shown and the equilibrium K d s obtained upon fitting of the raw data is mentioned in each panel. ( A, B ) Interaction of UP1 with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( C, D ) Interaction of UP1+RGG with the single stranded Tel22ss DNA in the presence of 100 mM NaCl and 100 mM KCl respectively. ( E, F ) Interaction of UP1 with the Na + and K + forms of Tel22 G-quadruplex DNA respectively. ( G, H ) Interaction of UP1+RGG with the Na + and K + forms of Tel22 G-quadruplex DNA respectively.

    Article Snippet: DNA preparation for CD, NMR and fluorescence kinetics experiments The DNA sequences (Table ) for binding studies with UP1, UP1+RGG and the RGG-box were ordered from Eurofins except the abasicloops-Tel22 DNA that was ordered from Dharmacon.

    Techniques:

    Relationship between the Illumina DNA-sequencing read depth and the copy number inferred by ddPCR. (A) The Y-axis represents copy numbers per μl inferred by ddPCR. Black circles (gray background) and open circles (black background) indicate three-copy genes and single-copy genes, respectively. All points and error bars represent averages of four replicates and 95% CIs. (B) Each dot represents an A. halleri gene. The X-axis represents the Illumina DNA-sequencing read depth, which is the number of reads per 1 Kbp per 1 million reads. The Y-axis represents copy numbers per μl, which were inferred by ddPCR. The regression line was calculated with the simple formula Y = αX; α was inferred by the least squares method.

    Journal: DNA Research: An International Journal for Rapid Publication of Reports on Genes and Genomes

    Article Title: Functional divergence of duplicate genes several million years after gene duplication in Arabidopsis

    doi: 10.1093/dnares/dsy005

    Figure Lengend Snippet: Relationship between the Illumina DNA-sequencing read depth and the copy number inferred by ddPCR. (A) The Y-axis represents copy numbers per μl inferred by ddPCR. Black circles (gray background) and open circles (black background) indicate three-copy genes and single-copy genes, respectively. All points and error bars represent averages of four replicates and 95% CIs. (B) Each dot represents an A. halleri gene. The X-axis represents the Illumina DNA-sequencing read depth, which is the number of reads per 1 Kbp per 1 million reads. The Y-axis represents copy numbers per μl, which were inferred by ddPCR. The regression line was calculated with the simple formula Y = αX; α was inferred by the least squares method.

    Article Snippet: We then inferred SSDs in a species after the divergence of closed species throughout the reading depth of Illumina DNA sequencing.

    Techniques: DNA Sequencing

    Three sets of OGGs among non- Arabidopsis species, A. thaliana , A. lyrata and A. halleri. OGGs between A. lyrata and A. halleri were defined as AL–AH OGGs. There were 25,833, 26,428 and 26,007 AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes, Illumina paired-end DNA-sequencing reads including pseudogene-like genes and the available A. halleri genome, respectively. OGGs among A. thaliana , A. lyrata and A. halleri were defined as AT–AL–AH OGGs. There were 22,105, 22,684 and 21,537 AT–AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes, Illumina paired-end DNA-sequencing reads including pseudogene-like genes and the available A. halleri genome, respectively. OGGs among non- Arabidopsis species ( B. rapa , B. stricta , C. grandiflora , C. rubella , E. salsugineum ), A. thaliana , A. lyrata and A. halleri were defined as nonA-AT–AL–AH OGGs. There were 17,669 and 17,925 non-A-AT–AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes and the available A. halleri genome, respectively.

    Journal: DNA Research: An International Journal for Rapid Publication of Reports on Genes and Genomes

    Article Title: Functional divergence of duplicate genes several million years after gene duplication in Arabidopsis

    doi: 10.1093/dnares/dsy005

    Figure Lengend Snippet: Three sets of OGGs among non- Arabidopsis species, A. thaliana , A. lyrata and A. halleri. OGGs between A. lyrata and A. halleri were defined as AL–AH OGGs. There were 25,833, 26,428 and 26,007 AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes, Illumina paired-end DNA-sequencing reads including pseudogene-like genes and the available A. halleri genome, respectively. OGGs among A. thaliana , A. lyrata and A. halleri were defined as AT–AL–AH OGGs. There were 22,105, 22,684 and 21,537 AT–AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes, Illumina paired-end DNA-sequencing reads including pseudogene-like genes and the available A. halleri genome, respectively. OGGs among non- Arabidopsis species ( B. rapa , B. stricta , C. grandiflora , C. rubella , E. salsugineum ), A. thaliana , A. lyrata and A. halleri were defined as nonA-AT–AL–AH OGGs. There were 17,669 and 17,925 non-A-AT–AL–AH OGGs based on Illumina paired-end DNA-sequencing reads without any pseudogene-like genes and the available A. halleri genome, respectively.

    Article Snippet: We then inferred SSDs in a species after the divergence of closed species throughout the reading depth of Illumina DNA sequencing.

    Techniques: DNA Sequencing

    Identification of minicircles in experimentally infected Beta vulgaris plants by the NGS approach. a Diagram of the bioinformatic pipeline used for the detection of minicircles. The tools used in each step are indicated in parentheses (refer to the Methods section for details). b Schematic representation of the eight BCTIV/ Beta vulgaris hybrid scaffolds assembled after the filtering step described in a . The junction coordinates referring to the BCTIV genome are indicated above each recombination point. The sizes (nt) of the B. vulgaris DNA fragments (blue portions) are reported on the right. c In vivo validation of the circular nature of the scaffolds reported in b , performed by inverse PCR (see also Supplementary Figure 3). The approximate positions of the primers specific for the non-viral portions of each minicircle (in blue) are indicated by arrows. DNA from mock-inoculated B. vulgaris . d Distribution of minicircle junctions between viral and host DNA. The proximal and distal junctions are marked with red and black arrows, respectively. BCTIV ORFs are represented by yellow boxes. e Relative contributions of B. vulgaris and BCTIV DNA to minicircles identified by NGS. f

    Journal: Nature Communications

    Article Title: Virus-mediated export of chromosomal DNA in plants

    doi: 10.1038/s41467-018-07775-w

    Figure Lengend Snippet: Identification of minicircles in experimentally infected Beta vulgaris plants by the NGS approach. a Diagram of the bioinformatic pipeline used for the detection of minicircles. The tools used in each step are indicated in parentheses (refer to the Methods section for details). b Schematic representation of the eight BCTIV/ Beta vulgaris hybrid scaffolds assembled after the filtering step described in a . The junction coordinates referring to the BCTIV genome are indicated above each recombination point. The sizes (nt) of the B. vulgaris DNA fragments (blue portions) are reported on the right. c In vivo validation of the circular nature of the scaffolds reported in b , performed by inverse PCR (see also Supplementary Figure 3). The approximate positions of the primers specific for the non-viral portions of each minicircle (in blue) are indicated by arrows. DNA from mock-inoculated B. vulgaris . d Distribution of minicircle junctions between viral and host DNA. The proximal and distal junctions are marked with red and black arrows, respectively. BCTIV ORFs are represented by yellow boxes. e Relative contributions of B. vulgaris and BCTIV DNA to minicircles identified by NGS. f

    Article Snippet: The genomic DNA sequencing libraries were prepared using the TruSeq DNA PCR-Free LT Library Prep Kit following the manufacturer’s instructions (Illumina, San Diego, CA), starting from 1.1 µg of sonicated DNA.

    Techniques: Infection, Next-Generation Sequencing, In Vivo, Inverse PCR

    The DNA sequencing of rs840088. Direct sequencing of a subgroup of samples with the same primers for PCR-RFLP was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an ABI PRISM 3100 DNA sequencer (Applied Biosystems, Foster City, USA). (A) GG genotype. (B) GA genotype. (C) AA genotype. 82×48 mm (300×300 DPI).

    Journal: Yonsei Medical Journal

    Article Title: The Association of SERPINE2 Gene with COPD in a Chinese Han Population

    doi: 10.3349/ymj.2011.52.6.953

    Figure Lengend Snippet: The DNA sequencing of rs840088. Direct sequencing of a subgroup of samples with the same primers for PCR-RFLP was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an ABI PRISM 3100 DNA sequencer (Applied Biosystems, Foster City, USA). (A) GG genotype. (B) GA genotype. (C) AA genotype. 82×48 mm (300×300 DPI).

    Article Snippet: Sequencing was performed using the BigDye Terminator v1.1 Cycle Sequencing kit (Applied Biosystems), and analyzed on an ABI PRISM 3100 DNA sequencer (Applied Biosystems, Foster City, CA, USA).

    Techniques: DNA Sequencing, Sequencing, Polymerase Chain Reaction