Structured Review

TaKaRa polymerase chain reaction pcr mixture
Amplified partial fragment of 16S rRNA gene from crow 1 (A), crow 2 (B), and crow 3 (C) for evaluation of intestinal microbiomes. Control <t>PCR</t> was performed under the same conditions except for without template <t>DNA</t> (D). Molecular size marker (M) shows 1.5, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, and 0.1 kb from top to bottom.
Polymerase Chain Reaction Pcr Mixture, supplied by TaKaRa, used in various techniques. Bioz Stars score: 91/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/polymerase chain reaction pcr mixture/product/TaKaRa
Average 91 stars, based on 31 article reviews
Price from $9.99 to $1999.99
polymerase chain reaction pcr mixture - by Bioz Stars, 2020-08
91/100 stars

Images

1) Product Images from "Population Abundance of Potentially Pathogenic Organisms in Intestinal Microbiome of Jungle Crow (Corvus macrorhynchos) Shown with 16S rRNA Gene-Based Microbial Community Analysis"

Article Title: Population Abundance of Potentially Pathogenic Organisms in Intestinal Microbiome of Jungle Crow (Corvus macrorhynchos) Shown with 16S rRNA Gene-Based Microbial Community Analysis

Journal: BioMed Research International

doi: 10.1155/2013/438956

Amplified partial fragment of 16S rRNA gene from crow 1 (A), crow 2 (B), and crow 3 (C) for evaluation of intestinal microbiomes. Control PCR was performed under the same conditions except for without template DNA (D). Molecular size marker (M) shows 1.5, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, and 0.1 kb from top to bottom.
Figure Legend Snippet: Amplified partial fragment of 16S rRNA gene from crow 1 (A), crow 2 (B), and crow 3 (C) for evaluation of intestinal microbiomes. Control PCR was performed under the same conditions except for without template DNA (D). Molecular size marker (M) shows 1.5, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, and 0.1 kb from top to bottom.

Techniques Used: Amplification, Polymerase Chain Reaction, Marker

Related Articles

Amplification:

Article Title: Non-Redundant and Overlapping Oncogenic Readouts of Non-Canonical and Novel Colorectal Cancer KRAS and NRAS Mutants
Article Snippet: .. The mutant KRAS and NRAS variant fragments were amplified in a polymerase chain reaction (PCR) mixture containing a final concentration of 1X PCR buffer (Titanium® Taq PCR buffer, Clontech Laboratories, Inc., Mountainview, CA, USA), 0.125 μM of each deoxynucleoside triphosphate (Promega, Madison, WI, USA), 2 μM each of the forward and reverse primers, 1U Taq polymerase (Titanium® Taq polymerase, Clontech Laboratories, Inc.), and 50 ng of the pTargeT-WT KRAS or pTargeT-WT NRAS template. .. Amplification was performed with an initial denaturation temperature of 94 °C for 5 min, followed by 25–30 cycles of denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, and extension at 72 °C for 30 s, with a final extension step at 72 °C for 10 min.

Mutagenesis:

Article Title: Non-Redundant and Overlapping Oncogenic Readouts of Non-Canonical and Novel Colorectal Cancer KRAS and NRAS Mutants
Article Snippet: .. The mutant KRAS and NRAS variant fragments were amplified in a polymerase chain reaction (PCR) mixture containing a final concentration of 1X PCR buffer (Titanium® Taq PCR buffer, Clontech Laboratories, Inc., Mountainview, CA, USA), 0.125 μM of each deoxynucleoside triphosphate (Promega, Madison, WI, USA), 2 μM each of the forward and reverse primers, 1U Taq polymerase (Titanium® Taq polymerase, Clontech Laboratories, Inc.), and 50 ng of the pTargeT-WT KRAS or pTargeT-WT NRAS template. .. Amplification was performed with an initial denaturation temperature of 94 °C for 5 min, followed by 25–30 cycles of denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, and extension at 72 °C for 30 s, with a final extension step at 72 °C for 10 min.

Isolation:

Article Title: Population Abundance of Potentially Pathogenic Organisms in Intestinal Microbiome of Jungle Crow (Corvus macrorhynchos) Shown with 16S rRNA Gene-Based Microbial Community Analysis
Article Snippet: .. PCR Conditions Twenty-five to fifty nanograms of isolated DNA were transferred to a PCR tube containing 50 μ L of a polymerase chain reaction (PCR) mixture containing LA Taq DNA polymerase (TaKaRa Bio, Shiga, Japan). .. For DNA evaluation, the eubacterial 16S rRNA gene sequence intervened between UNIV341F (5′-CCTACGGGAGGCAGCAG-3′) and UNIV907R (5′-CCCCGTCAATTCCTTTGAGTTT-3′) was amplified.

Concentration Assay:

Article Title: Non-Redundant and Overlapping Oncogenic Readouts of Non-Canonical and Novel Colorectal Cancer KRAS and NRAS Mutants
Article Snippet: .. The mutant KRAS and NRAS variant fragments were amplified in a polymerase chain reaction (PCR) mixture containing a final concentration of 1X PCR buffer (Titanium® Taq PCR buffer, Clontech Laboratories, Inc., Mountainview, CA, USA), 0.125 μM of each deoxynucleoside triphosphate (Promega, Madison, WI, USA), 2 μM each of the forward and reverse primers, 1U Taq polymerase (Titanium® Taq polymerase, Clontech Laboratories, Inc.), and 50 ng of the pTargeT-WT KRAS or pTargeT-WT NRAS template. .. Amplification was performed with an initial denaturation temperature of 94 °C for 5 min, followed by 25–30 cycles of denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, and extension at 72 °C for 30 s, with a final extension step at 72 °C for 10 min.

Polymerase Chain Reaction:

Article Title: Evidence of Subterranean Termite Feeding Deterrent Produced by Brown Rot Fungus Fibroporia radiculosa (Peck) Parmasto 1968 (Polyporales, Fomitopsidaceae)
Article Snippet: .. The polymerase chain reaction (PCR) mixture was prepared using TaKaRa Ex Taq (TaKaRa Bio, Shiga, Japan), and primers for ITS1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS4 (5’-TCCTCCGCTTATTGATATGC) were added. .. PCR was performed with a 2720 Thermal Cycler (Applied Biosystems, Foster City, CA, USA) with the following parameters: initial denaturation for 3 min at 94 °C; 25 cycles of denaturation for 15 s at 94 °C, annealing for 30 s at 55 °C, and extension for 30 s at 72 °C; and a final extension for 5 min at 72 °C.

Article Title: Population Abundance of Potentially Pathogenic Organisms in Intestinal Microbiome of Jungle Crow (Corvus macrorhynchos) Shown with 16S rRNA Gene-Based Microbial Community Analysis
Article Snippet: .. PCR Conditions Twenty-five to fifty nanograms of isolated DNA were transferred to a PCR tube containing 50 μ L of a polymerase chain reaction (PCR) mixture containing LA Taq DNA polymerase (TaKaRa Bio, Shiga, Japan). .. For DNA evaluation, the eubacterial 16S rRNA gene sequence intervened between UNIV341F (5′-CCTACGGGAGGCAGCAG-3′) and UNIV907R (5′-CCCCGTCAATTCCTTTGAGTTT-3′) was amplified.

Article Title: Taxonomy of myid bivalves from fragmented brackish-water habitats in India, with a description of a new genus Indosphenia ( Myidae, Myoidea, Myidae)
Article Snippet: .. The polymerase chain reaction ( PCR ) mixture consisted of 25 μL Master Mix (Takara Clontech EmeraldAmp® GT PCR Master Mix), 1 μL forward primer, 1 μL reverse primer, 8 μL template DNA, and 15 μL distilled deionised water. .. The amplification primers were LCO-1490 F (5'- GGTCAACAAATCATAAAGATATTGG-3') and HCO-2198 R (5'-TAAACTTCAGGGTGACCAAAAAATCA-3'), used for amplifying mitochondrial cytochrome c oxidase subunit I ( mtCOI ) gene sequences ( ).

Article Title: Feasibility and effect of ultrasound microbubble-mediated wild-type p53 gene transfection of HeLa cells
Article Snippet: .. Polymerase chain reaction (PCR) mixture was from Takara Biotechnology Co., Ltd., (Dalian, China). .. Lipofectamine™ 2000 reagent was obtained from Invitrogen (Carlsbad, CA, USA).

Article Title: Non-Redundant and Overlapping Oncogenic Readouts of Non-Canonical and Novel Colorectal Cancer KRAS and NRAS Mutants
Article Snippet: .. The mutant KRAS and NRAS variant fragments were amplified in a polymerase chain reaction (PCR) mixture containing a final concentration of 1X PCR buffer (Titanium® Taq PCR buffer, Clontech Laboratories, Inc., Mountainview, CA, USA), 0.125 μM of each deoxynucleoside triphosphate (Promega, Madison, WI, USA), 2 μM each of the forward and reverse primers, 1U Taq polymerase (Titanium® Taq polymerase, Clontech Laboratories, Inc.), and 50 ng of the pTargeT-WT KRAS or pTargeT-WT NRAS template. .. Amplification was performed with an initial denaturation temperature of 94 °C for 5 min, followed by 25–30 cycles of denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, and extension at 72 °C for 30 s, with a final extension step at 72 °C for 10 min.

Article Title: Isolation and 2,4-D-degrading characteristics of Cupriaviduscampinensis BJ71
Article Snippet: .. Each polymerase chain reaction (PCR) mixture contained 1 × Premix Taq (Ex Taq Version, Takara Bio, Shiga, Japan), 0.4 μM each primer, and ~50 ng template DNA. .. The PCR amplification conditions were as follows: 95 °C for 5 min; followed by 35 cycles of 95 °C for 45 s, 55 °C for 45 s, and 72 °C for 90 s. To ensure complete elongation, a final step of 72 °C for 7 min was performed.

Article Title: Effects of Dietary Supplementation With Enterococcus faecium and Clostridium butyricum, Either Alone or in Combination, on Growth and Fecal Microbiota Composition of Post-weaning Pigs at a Commercial Farm
Article Snippet: .. Each 50 μL of polymerase chain reaction (PCR) mixture contained 1 μL of sample DNA, 21 μL of MilliQ® water, 25 μL of 2X MightyAmp® Buffer, 1 μL of forward primer (5 μM), 1 μL of reverse primer (5 μM), and 1 μL (1.25 U) of MightyAmp DNA Polymerase (Takara, Tokyo, Japan). ..

Article Title: Use of RNAi technology to develop a PRSV-resistant transgenic papaya
Article Snippet: .. The polymerase chain reaction (PCR) mixture contained 1 µl of cDNA, 0.5 µl of each primer (10 µM), 2 µl of 10 × PCR buffer, 1 µl of dNTP, 0.25 U of Taq DNA polymerase (Takara), and nuclease-free water to a final volume of 20 µl. .. The amplification consisted of 35 cycles at 94 °C (30 s), 50 °C (1 min), and 72 °C (1 min); the initial cycle incorporated a melting step of 94 °C for 3 min and the final cycle a synthesis step of 72 °C for 10 min. PCR products were electrophoresed on 1.5% agarose gel, and visualized after ethidium bromide staining.

Variant Assay:

Article Title: Non-Redundant and Overlapping Oncogenic Readouts of Non-Canonical and Novel Colorectal Cancer KRAS and NRAS Mutants
Article Snippet: .. The mutant KRAS and NRAS variant fragments were amplified in a polymerase chain reaction (PCR) mixture containing a final concentration of 1X PCR buffer (Titanium® Taq PCR buffer, Clontech Laboratories, Inc., Mountainview, CA, USA), 0.125 μM of each deoxynucleoside triphosphate (Promega, Madison, WI, USA), 2 μM each of the forward and reverse primers, 1U Taq polymerase (Titanium® Taq polymerase, Clontech Laboratories, Inc.), and 50 ng of the pTargeT-WT KRAS or pTargeT-WT NRAS template. .. Amplification was performed with an initial denaturation temperature of 94 °C for 5 min, followed by 25–30 cycles of denaturation at 94 °C for 30 s, annealing at 55 °C for 30 s, and extension at 72 °C for 30 s, with a final extension step at 72 °C for 10 min.

Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 93
    TaKaRa tb green pcr master mix
    A , Quantitative <t>PCR</t> (qPCR) analysis of DONSON mRNA amount in HeLa cells. Total RNA was isolated from HeLa cells after 48 h treatment of the indicated siRNAs, and was subjected to reverse transcription. The resulting <t>cDNA</t> was subjected to real-time qPCR analysis. B, Time-lapse observation of DONSON localization at different cell cycle stages. HeLa GFP-DONSON cells were visualized every 10 min for 24 h after 24 h treatment of 1 μg/mL Doxycycline and 3 h treatment of 100 nM SiR-DNA. DNA (blue) and GFP-DONSON (green). Arrowheads indicate GFP-DONSON foci. C, The centrosomal localization of DONSON in mitosis. HeLa GFP-DONSON cells were fixed after 24 h treatment of 1 μg/mL Doxycycline and were immunostained with antibodies against GFP (green) and CP110 (red). D, Phenotype rescue experiments by expression of RNAi-resistant (RNAi-R) form of full-length (FL) or nuclear localization signal deleted (ΔNLS) DONSON. HeLa cells were treated with siControl or siDONSON, followed by transfection with FLAG empty (control), RNAi-resistant DONSON-FL, or DONSON-ΔNLS. The cells were immunostained with antibodies against FLAG (green) and Cep192 (red). Arrowheads indicate Cep192 foci. E, Histograms represent frequency of interphase cells with the indicated phenotypes observed in D. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). Tukey’s multiple comparisons test was used in E to obtain P value. **, p
    Tb Green Pcr Master Mix, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/tb green pcr master mix/product/TaKaRa
    Average 93 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    tb green pcr master mix - by Bioz Stars, 2020-08
    93/100 stars
      Buy from Supplier

    89
    TaKaRa terra pcr direct polymerase mix
    Generation of Ush2a null alleles in OC-k1 cells. (a) <t>RT-PCR</t> of Ush2a , Whrn ( Ush2d ) and Adgrv1 ( Ush2c ) genes in OC-k1 cells. Ush2a RT-serves as experimental control (b) Immunostaining of OC-k1 cells with anti-Ush2a and anti-acetylated alpha tubulin antibodies. Expression of Ush2a (green) was detected at the base of primary cilia (red) in OC-k1 cells. Scale bar= 5μm. (c) Schematic of the <t>sgRNA</t> targets on mouse Ush2a gene. The targeted genomic sites are indicated in green (for exon3 sg-mEX3), cyan (for exon 5 – sg-mEX5) and the protospacer adjacent motif (PAM) sequence is marked in red. (d) T7 endonuclease cleavage assay performed on pooled flow-sorted GFP positive OC-k1 cells revealed indel formation at the target locus of sgRNAs on exon 3 and 5. * represents the cleaved DNA fragments. (e) Indel rate (% of edited reads out of total reads) detected by NGS deep sequencing of sgRNA target sites on exon 3 and exon 5 from the pooled GFP positive OC-k1 cells. The distribution of in-frame and out-of-frame indels from sg-mEX5 targeted cells is shown in the pie chart on the right. (f) T7E1 assay for single sorted clones revealed cleaved DNA fragments (*) in single sorted clone D, F and J. (g) Sequence patterns and rates of editing event at the target site of Ush2a locus in clones D, F, J and wild type OC-k1 cells by NGS deep sequencing analysis. The percentage of in-frame and out-of-frame alleles in clones D, F and J is shown in the right most column. All the Ush2a alleles in clone J are out-of-frame, which makes it a null line for Ush2a gene.
    Terra Pcr Direct Polymerase Mix, supplied by TaKaRa, used in various techniques. Bioz Stars score: 89/100, based on 33 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/terra pcr direct polymerase mix/product/TaKaRa
    Average 89 stars, based on 33 article reviews
    Price from $9.99 to $1999.99
    terra pcr direct polymerase mix - by Bioz Stars, 2020-08
    89/100 stars
      Buy from Supplier

    90
    TaKaRa qrt pcr mixture
    Overexpression of PpGLK1 in u/u tomato. (A) <t>qRT-PCR</t> and western blot analysis confirming PpGLK1 overexpression in u/u tomato. (B) Phenotype of PpGLK1 -overexpressing u/u tomato. (C) Chlorophyll contents in u/u tomato and PpGLK1 -overexpressing u/u tomato. (D) Chloroplast development in fruit observed by TEM in PpGLK1 -overexpressing u/u tomato fruit and u/u tomato fruit. Data is presented as the means ± SD, n = 3. Different letters above bars denote statistical significance according to one-way ANOVA and Duncan's test ( P
    Qrt Pcr Mixture, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/qrt pcr mixture/product/TaKaRa
    Average 90 stars, based on 28 article reviews
    Price from $9.99 to $1999.99
    qrt pcr mixture - by Bioz Stars, 2020-08
    90/100 stars
      Buy from Supplier

    94
    TaKaRa dna template
    Inverse <t>PCR</t> (iPCR) and sequencing of the boundary region surrounding the Mk + 1.3 locus. ( a ) Agarose gel electrophoresis of nested PCR products stained with ethidium bromide. The gel is the full length gel and lanes have not been cropped or stitched. ( b ) Chromatograms generated from direct sequencing of the iPCR products. Black and red letters above the chromatograms indicate the sequences of chromosomal positions chr 7: 62,416,891–62,416,891 and chr 7: 62,268,615–62,268,638 (on the University of California, Santa Cruz, genome browser (GRCm38/mm10)), respectively. W: wild-type <t>DNA,</t> Tg: hemizygous transgenic DNA, M: molecular size marker.
    Dna Template, supplied by TaKaRa, used in various techniques. Bioz Stars score: 94/100, based on 528 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dna template/product/TaKaRa
    Average 94 stars, based on 528 article reviews
    Price from $9.99 to $1999.99
    dna template - by Bioz Stars, 2020-08
    94/100 stars
      Buy from Supplier

    Image Search Results


    A , Quantitative PCR (qPCR) analysis of DONSON mRNA amount in HeLa cells. Total RNA was isolated from HeLa cells after 48 h treatment of the indicated siRNAs, and was subjected to reverse transcription. The resulting cDNA was subjected to real-time qPCR analysis. B, Time-lapse observation of DONSON localization at different cell cycle stages. HeLa GFP-DONSON cells were visualized every 10 min for 24 h after 24 h treatment of 1 μg/mL Doxycycline and 3 h treatment of 100 nM SiR-DNA. DNA (blue) and GFP-DONSON (green). Arrowheads indicate GFP-DONSON foci. C, The centrosomal localization of DONSON in mitosis. HeLa GFP-DONSON cells were fixed after 24 h treatment of 1 μg/mL Doxycycline and were immunostained with antibodies against GFP (green) and CP110 (red). D, Phenotype rescue experiments by expression of RNAi-resistant (RNAi-R) form of full-length (FL) or nuclear localization signal deleted (ΔNLS) DONSON. HeLa cells were treated with siControl or siDONSON, followed by transfection with FLAG empty (control), RNAi-resistant DONSON-FL, or DONSON-ΔNLS. The cells were immunostained with antibodies against FLAG (green) and Cep192 (red). Arrowheads indicate Cep192 foci. E, Histograms represent frequency of interphase cells with the indicated phenotypes observed in D. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). Tukey’s multiple comparisons test was used in E to obtain P value. **, p

    Journal: bioRxiv

    Article Title: DONSON, a gene responsible for microcephalic primordial dwarfism, ensures proper centriole duplication cycle by maintaining centriole engagement during interphase

    doi: 10.1101/2020.05.10.086777

    Figure Lengend Snippet: A , Quantitative PCR (qPCR) analysis of DONSON mRNA amount in HeLa cells. Total RNA was isolated from HeLa cells after 48 h treatment of the indicated siRNAs, and was subjected to reverse transcription. The resulting cDNA was subjected to real-time qPCR analysis. B, Time-lapse observation of DONSON localization at different cell cycle stages. HeLa GFP-DONSON cells were visualized every 10 min for 24 h after 24 h treatment of 1 μg/mL Doxycycline and 3 h treatment of 100 nM SiR-DNA. DNA (blue) and GFP-DONSON (green). Arrowheads indicate GFP-DONSON foci. C, The centrosomal localization of DONSON in mitosis. HeLa GFP-DONSON cells were fixed after 24 h treatment of 1 μg/mL Doxycycline and were immunostained with antibodies against GFP (green) and CP110 (red). D, Phenotype rescue experiments by expression of RNAi-resistant (RNAi-R) form of full-length (FL) or nuclear localization signal deleted (ΔNLS) DONSON. HeLa cells were treated with siControl or siDONSON, followed by transfection with FLAG empty (control), RNAi-resistant DONSON-FL, or DONSON-ΔNLS. The cells were immunostained with antibodies against FLAG (green) and Cep192 (red). Arrowheads indicate Cep192 foci. E, Histograms represent frequency of interphase cells with the indicated phenotypes observed in D. Values are mean percentages ± s.d. from three independent experiments (n = 30 for each experiment). Tukey’s multiple comparisons test was used in E to obtain P value. **, p

    Article Snippet: Reverse transcription and real-time qPCR Total RNA (1 μg) isolated from cells with the use of TRIZOL reagent (Thermo Fischer Scientific) was subject to reverse transcription using Quantiscript Reverse Transcription Kit (QIAGEN), and the resulting cDNA was subjected to real-time qPCR analysis with TB Green PCR Master Mix (TaKaRa) and specific primers in a StepOnePlus Real-Time PCR system (Applied Biosystems).

    Techniques: Real-time Polymerase Chain Reaction, Isolation, Expressing, Transfection

    Generation of Ush2a null alleles in OC-k1 cells. (a) RT-PCR of Ush2a , Whrn ( Ush2d ) and Adgrv1 ( Ush2c ) genes in OC-k1 cells. Ush2a RT-serves as experimental control (b) Immunostaining of OC-k1 cells with anti-Ush2a and anti-acetylated alpha tubulin antibodies. Expression of Ush2a (green) was detected at the base of primary cilia (red) in OC-k1 cells. Scale bar= 5μm. (c) Schematic of the sgRNA targets on mouse Ush2a gene. The targeted genomic sites are indicated in green (for exon3 sg-mEX3), cyan (for exon 5 – sg-mEX5) and the protospacer adjacent motif (PAM) sequence is marked in red. (d) T7 endonuclease cleavage assay performed on pooled flow-sorted GFP positive OC-k1 cells revealed indel formation at the target locus of sgRNAs on exon 3 and 5. * represents the cleaved DNA fragments. (e) Indel rate (% of edited reads out of total reads) detected by NGS deep sequencing of sgRNA target sites on exon 3 and exon 5 from the pooled GFP positive OC-k1 cells. The distribution of in-frame and out-of-frame indels from sg-mEX5 targeted cells is shown in the pie chart on the right. (f) T7E1 assay for single sorted clones revealed cleaved DNA fragments (*) in single sorted clone D, F and J. (g) Sequence patterns and rates of editing event at the target site of Ush2a locus in clones D, F, J and wild type OC-k1 cells by NGS deep sequencing analysis. The percentage of in-frame and out-of-frame alleles in clones D, F and J is shown in the right most column. All the Ush2a alleles in clone J are out-of-frame, which makes it a null line for Ush2a gene.

    Journal: bioRxiv

    Article Title: Exon 13-skipped USH2A protein retains functional integrity in mice, suggesting an exo-skipping therapeutic approach to treat USH2A-associated disease

    doi: 10.1101/2020.02.04.934240

    Figure Lengend Snippet: Generation of Ush2a null alleles in OC-k1 cells. (a) RT-PCR of Ush2a , Whrn ( Ush2d ) and Adgrv1 ( Ush2c ) genes in OC-k1 cells. Ush2a RT-serves as experimental control (b) Immunostaining of OC-k1 cells with anti-Ush2a and anti-acetylated alpha tubulin antibodies. Expression of Ush2a (green) was detected at the base of primary cilia (red) in OC-k1 cells. Scale bar= 5μm. (c) Schematic of the sgRNA targets on mouse Ush2a gene. The targeted genomic sites are indicated in green (for exon3 sg-mEX3), cyan (for exon 5 – sg-mEX5) and the protospacer adjacent motif (PAM) sequence is marked in red. (d) T7 endonuclease cleavage assay performed on pooled flow-sorted GFP positive OC-k1 cells revealed indel formation at the target locus of sgRNAs on exon 3 and 5. * represents the cleaved DNA fragments. (e) Indel rate (% of edited reads out of total reads) detected by NGS deep sequencing of sgRNA target sites on exon 3 and exon 5 from the pooled GFP positive OC-k1 cells. The distribution of in-frame and out-of-frame indels from sg-mEX5 targeted cells is shown in the pie chart on the right. (f) T7E1 assay for single sorted clones revealed cleaved DNA fragments (*) in single sorted clone D, F and J. (g) Sequence patterns and rates of editing event at the target site of Ush2a locus in clones D, F, J and wild type OC-k1 cells by NGS deep sequencing analysis. The percentage of in-frame and out-of-frame alleles in clones D, F and J is shown in the right most column. All the Ush2a alleles in clone J are out-of-frame, which makes it a null line for Ush2a gene.

    Article Snippet: The sgRNA target locus on exon 3 or 5 was PCR amplified with Terra™ PCR Direct Polymerase Mix (Takara) for 35 cycles using primer sets mEX3F and mEX3R or mEX5F and mEX5R ( ).

    Techniques: Reverse Transcription Polymerase Chain Reaction, Immunostaining, Expressing, Sequencing, Cleavage Assay, Next-Generation Sequencing, Clone Assay

    Overexpression of PpGLK1 in u/u tomato. (A) qRT-PCR and western blot analysis confirming PpGLK1 overexpression in u/u tomato. (B) Phenotype of PpGLK1 -overexpressing u/u tomato. (C) Chlorophyll contents in u/u tomato and PpGLK1 -overexpressing u/u tomato. (D) Chloroplast development in fruit observed by TEM in PpGLK1 -overexpressing u/u tomato fruit and u/u tomato fruit. Data is presented as the means ± SD, n = 3. Different letters above bars denote statistical significance according to one-way ANOVA and Duncan's test ( P

    Journal: Frontiers in Plant Science

    Article Title: Transcriptomic and Functional Analyses Reveal That PpGLK1 Regulates Chloroplast Development in Peach (Prunus persica)

    doi: 10.3389/fpls.2018.00034

    Figure Lengend Snippet: Overexpression of PpGLK1 in u/u tomato. (A) qRT-PCR and western blot analysis confirming PpGLK1 overexpression in u/u tomato. (B) Phenotype of PpGLK1 -overexpressing u/u tomato. (C) Chlorophyll contents in u/u tomato and PpGLK1 -overexpressing u/u tomato. (D) Chloroplast development in fruit observed by TEM in PpGLK1 -overexpressing u/u tomato fruit and u/u tomato fruit. Data is presented as the means ± SD, n = 3. Different letters above bars denote statistical significance according to one-way ANOVA and Duncan's test ( P

    Article Snippet: The qRT-PCR mixture consisted of 10 μL SYBR Premix Ex Taq (TaKaRa Biotechnology, Dalian, China), 1 μL cDNA, and 1 μL each primer pair.

    Techniques: Over Expression, Quantitative RT-PCR, Western Blot, Transmission Electron Microscopy

    Inverse PCR (iPCR) and sequencing of the boundary region surrounding the Mk + 1.3 locus. ( a ) Agarose gel electrophoresis of nested PCR products stained with ethidium bromide. The gel is the full length gel and lanes have not been cropped or stitched. ( b ) Chromatograms generated from direct sequencing of the iPCR products. Black and red letters above the chromatograms indicate the sequences of chromosomal positions chr 7: 62,416,891–62,416,891 and chr 7: 62,268,615–62,268,638 (on the University of California, Santa Cruz, genome browser (GRCm38/mm10)), respectively. W: wild-type DNA, Tg: hemizygous transgenic DNA, M: molecular size marker.

    Journal: Scientific Reports

    Article Title: Application of droplet digital PCR in the analysis of genome integration and organization of the transgene in BAC transgenic mice

    doi: 10.1038/s41598-018-25001-x

    Figure Lengend Snippet: Inverse PCR (iPCR) and sequencing of the boundary region surrounding the Mk + 1.3 locus. ( a ) Agarose gel electrophoresis of nested PCR products stained with ethidium bromide. The gel is the full length gel and lanes have not been cropped or stitched. ( b ) Chromatograms generated from direct sequencing of the iPCR products. Black and red letters above the chromatograms indicate the sequences of chromosomal positions chr 7: 62,416,891–62,416,891 and chr 7: 62,268,615–62,268,638 (on the University of California, Santa Cruz, genome browser (GRCm38/mm10)), respectively. W: wild-type DNA, Tg: hemizygous transgenic DNA, M: molecular size marker.

    Article Snippet: Nested amplification around the circular DNA using primers oriented in a direction opposite to that of conventional PCR was performed using reaction mixtures containing DNA template, 1 × Ex Taq buffer (containing 2.0 mM of MgCl2 ), 250 μM of dNTP mix, 0.5 μM of each primer, and 0.25 U/μl of Ex Taq DNA Polymerase (Takara Bio, Otsu, Japan).

    Techniques: Inverse PCR, Sequencing, Agarose Gel Electrophoresis, Nested PCR, Staining, Generated, Transgenic Assay, Marker