terra pcr direct polymerase mix  (TaKaRa)

 
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    Name:
    Terra PCR Direct Polymerase Mix
    Description:

    Catalog Number:
    639271
    Price:
    None
    Score:
    85
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    Structured Review

    TaKaRa terra pcr direct polymerase mix
    Targeted deletion of GH receptor in MΦ . A , genomic <t>DNA</t> <t>PCR</t> for Cre and the floxed GH receptor exon 4; absence of AC band containing exon 4 in peritoneal MΦ with retention of band in liver DNA. B , GHR mRNA expression is extinguished in

    https://www.bioz.com/result/terra pcr direct polymerase mix/product/TaKaRa
    Average 99 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    terra pcr direct polymerase mix - by Bioz Stars, 2019-10
    99/100 stars

    Images

    1) Product Images from "Targeted Deletion of Growth Hormone (GH) Receptor in Macrophage Reveals Novel Osteopontin-mediated Effects of GH on Glucose Homeostasis and Insulin Sensitivity in Diet-induced Obesity"

    Article Title: Targeted Deletion of Growth Hormone (GH) Receptor in Macrophage Reveals Novel Osteopontin-mediated Effects of GH on Glucose Homeostasis and Insulin Sensitivity in Diet-induced Obesity

    Journal:

    doi: 10.1074/jbc.M113.460212

    Targeted deletion of GH receptor in MΦ . A , genomic DNA PCR for Cre and the floxed GH receptor exon 4; absence of AC band containing exon 4 in peritoneal MΦ with retention of band in liver DNA. B , GHR mRNA expression is extinguished in
    Figure Legend Snippet: Targeted deletion of GH receptor in MΦ . A , genomic DNA PCR for Cre and the floxed GH receptor exon 4; absence of AC band containing exon 4 in peritoneal MΦ with retention of band in liver DNA. B , GHR mRNA expression is extinguished in

    Techniques Used: Polymerase Chain Reaction, Expressing

    Related Articles

    Clone Assay:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: Paragraph title: DNA cloning method ... Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water.

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. PCR using genomic DNA extracted from Tg26 rat blood samples was performed in one step using LTR F and nested Gag R primers.

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: HepG2 cell line of hepatoblastoma and E. coli strains DH5α were conserved in our laboratory. .. Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Centrifugation:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: After centrifugation, the supernatant having the DNA template was used for PCR reactions. .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water.

    Amplification:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: The nuclear ITS1 DNA was amplified by primers Betta_ITS1_F2 and Betta_ITS1_R2, Betta_ITS1_F3 and Betta_ITS1_R3, or Betta_ITS_F2 and Betta_ITS_R4 designed by Mahidol University (see ). .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. The PCR conditions were 98 °C for 2 min followed by 35 cycles of 98 °C for 10 s, annealing at 52 °C for 15 s and elongation at 68 °C for 1 min/kb.

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: The specificity and sensitivity of the PCR and potential interference by inhibitors in the feces were determined before assay application [ ]. .. Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. DNA fragments were amplified using the following optimized thermos-cycling conditions: 95 °C/5 min for denaturation; 35 cycles of 94 °C /30 s, 55 °C /30 s, 72 °C /40 s; and 72 °C /10 min extension.

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: The specificity and sensitivity of the PCR and potential interference by inhibitors in the feces were determined before assay application [ ]. .. Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × ExTaq Buffer, MgSO4 25 mM, 0.25 μl ExTaq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. DNA fragments were amplified using the following optimized thermos-cycling conditions: 95 °C/5 min for denaturation; 35 cycles of 94 °C /30 s, 55 °C /30 s, 72 °C /40 s; and 72 °C /10 min extension.

    Article Title: Whole-Genome Re-Alignment Facilitates Development of Specific Molecular Markers for Races 1 and 4 of Xanthomonas campestris pv. campestris, the Cause of Black Rot Disease in Brassica oleracea
    Article Snippet: The race 4 specific primers were: Xcc1_46R4 designed between 1843518 and 1843057 and Xcc1_46R4 primer designed between 1842956 to 1842379; these two primers were expected to amplify 462 and 578 base pairs, respectively ( ). .. Emerald PCR master mix (Takara, Shiga, Japan) was used in Polymerase Chain Reaction (PCR) for amplification of the target regions with respective markers. .. A 20.0 μL of PCR reaction mixture containing forward and reverse primers (1.0 μL each), Emerald PCR master mix (9.0 μL), ultra-pure water (8 μL) and 1.0 μL DNA was used for PCR amplification.

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Transgenic seedlings were selected on MS agar medium containing hygromycin (25 μg/ml). .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. The T2 progeny of three independent transgenic lines were used for functional studies.

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: PCR amplification of the corresponding genomic region surrounding each SNP locus was performed in a TaKaRa PCR thermal cycler (TaKaRa TP600, China). .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: Paragraph title: DNA Extraction and PCR Amplification ... A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA).

    Article Title: Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity
    Article Snippet: Paragraph title: Performance improvements of whole-transcript amplification ... The use of this DNA polymerase (MightyAmp DNA Polymerase (Takara Bio, Inc., Tokyo, Japan), also marketed as Terra PCR Direct Polymerase (Clontech, Mountain View, CA, USA)) improved the yield of cDNA (see Additional file , Figure S2c and Figure S5) and the reproducibility of the WTA replication (see Additional file , Figure S2d and Figure S5).

    Article Title: Changing Patterns of Malaria Epidemiology between 2002 and 2010 in Western Kenya: The Fall and Rise of Malaria
    Article Snippet: From 2006 onward, only legs were used for PCR species identification. .. About one-third of the leg samples were analyzed by DNA extraction and amplification of the ribosomal DNA using the Terra-Direct PCR method (Clonetech, Mountain View, CA. .. The rest of the samples from 2006 to 2010 were analyzed by using Fast Tissue-to-PCR kit (Fermentas, Glen Burnie, MA) and individual specimen was identified by the standard PCR procedure .

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    Mass Spectrometry:

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Transgenic seedlings were selected on MS agar medium containing hygromycin (25 μg/ml). .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product.

    Synthesized:

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: First strand cDNAs were synthesized from 2.5 μg total RNA using the M-MLV reverse transcriptase kit (Transgen biotech, Beijing). .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Real-time Polymerase Chain Reaction:

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Both RT-PCR and qRT-PCR were used to analyze the expression level of REL2 /Os10g0562700 in WT and rel2 mutant. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions. .. For analysis of REL2 various tissues expression, a specific fragment was amplified by using primers 5’-TGATCATCGTGACTTCACAGGC-3’ and 5’-TCTACCAGACCACGGACTTGC-3’.

    Article Title: Mutation of kri1l causes definitive hematopoiesis failure via PERK-dependent excessive autophagy induction
    Article Snippet: RNA was reverse-transcribed using random hexamers and SuperScript III Reverse Transcriptase (Invitrogen). .. 2× PCR Mix (TaKaRa, Premix Ex Taq) containing SYBR Green I was used for the real-time quantitative PCR analysis with the Applied Biosystems 7900HT Fast Real-Time PCR System. .. The relative expression values were normalized against the internal control actin (QPCR primer sequences were listed in ).

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: Paragraph title: PCR and qPCR ... The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng).

    Article Title: Control of alternative end joining by the chromatin remodeler p400 ATPase
    Article Snippet: After RNaseH treatment (20 units), genomic DNA sample was digested or not with 20 units of restriction enzymes (BanI) at 37°C overnight. .. Two microliters of digested or not samples (20 ng) were used as templates in 25 μl of qPCR reaction containing 12.5 μl TAKARA Mix PCR. .. The percentage of ssDNA (ssDNA%) generated by resection at selected sites was determined as previously described ( ).

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: All ChIP primers used in the standard PCR and quantitative real-time PCR experiment are listed in Table . .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C.

    Quantitative RT-PCR:

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Both RT-PCR and qRT-PCR were used to analyze the expression level of REL2 /Os10g0562700 in WT and rel2 mutant. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions. .. For analysis of REL2 various tissues expression, a specific fragment was amplified by using primers 5’-TGATCATCGTGACTTCACAGGC-3’ and 5’-TCTACCAGACCACGGACTTGC-3’.

    SYBR Green Assay:

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Both RT-PCR and qRT-PCR were used to analyze the expression level of REL2 /Os10g0562700 in WT and rel2 mutant. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions. .. For analysis of REL2 various tissues expression, a specific fragment was amplified by using primers 5’-TGATCATCGTGACTTCACAGGC-3’ and 5’-TCTACCAGACCACGGACTTGC-3’.

    Article Title: Mutation of kri1l causes definitive hematopoiesis failure via PERK-dependent excessive autophagy induction
    Article Snippet: RNA was reverse-transcribed using random hexamers and SuperScript III Reverse Transcriptase (Invitrogen). .. 2× PCR Mix (TaKaRa, Premix Ex Taq) containing SYBR Green I was used for the real-time quantitative PCR analysis with the Applied Biosystems 7900HT Fast Real-Time PCR System. .. The relative expression values were normalized against the internal control actin (QPCR primer sequences were listed in ).

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C.

    Incubation:

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 1.5 µl purified RNA was mixed with 5 µl reaction mix (1× PCR buffer (Roche), 1.5 mM MgCl2 , 50 µM dNTPs, 2 ng/µl poly–T primer (TATAGAATTCGCGGCCGCTCGCGATTTTTTTTTTTTTTTTTTTTTTTT), 0.04 U/µl RNase inhibitor (Qiagen), 0.4 U/µl recombinant RNasin (Promega)). .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min. .. The PCR product was purified by gel purification, and 50 ng of the purified product was used for library preparation with Nextera DNA Sample Prep kits (Illumina).

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA. .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: Fecal DNA (6 from baseline, 12 per treatment) were isolated from approximately 100 mg samples using a commercially available kit (Mag-Bind Soil DNA Kit; Omega Bio-tek, Inc., Norcross, GA) according to the manufacturer’s protocol with the following modifications: the raw sample was transferred to a 2 mL sterile Safe-Lock tube (Eppendorf, Hauppauge, NY) with 0.3 g of acid washed beads (Scientific Asset Management, Basking Ridge, NJ) and 300 µL of SLX-Mlus Buffer, followed by bead-beating at frequency of 20 for 10 min (QIAGEN Inc., Valencia, CA); after samples were mixed with RNase A and DS Buffer, samples were incubated in a 90 °C water bath for 8 min and occasionally vortexed; the samples were centrifuged at a speed of 5,000 × g for 10 min and the supernatant was transferred; finally, DNA was eluted with 130 µL of elution buffer at the last step. .. A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA).

    Article Title: Sensitive and powerful single-cell RNA sequencing using mcSCRB-seq
    Article Snippet: Before library preparation, proteins were digested by incubation at 50 °C for 10 min. Proteinase K was then heat inactivated for 10 min at 80 °C. .. After heat inactivation for 10 min at 80 °C, 30 µl PCR master mix consisting of 1.25 U Terra direct polymerase (Clontech) 1.66 × Terra direct buffer and 0.33 µM SINGV6 primer (IDT) was added.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: All ChIP primers used in the standard PCR and quantitative real-time PCR experiment are listed in Table . .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    Expressing:

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min. .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min.

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′). .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Paragraph title: RNA Isolation and Gene Expression Analysis ... The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: HepG2 cell line of hepatoblastoma and E. coli strains DH5α were conserved in our laboratory. .. Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Modification:

    Article Title: Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity
    Article Snippet: In the third improvement point, to shorten the length of the remaining primers modified by the terminal transferase and thus make them targets for suppression PCR, we restricted the reaction time of the terminal transferase. .. The use of this DNA polymerase (MightyAmp DNA Polymerase (Takara Bio, Inc., Tokyo, Japan), also marketed as Terra PCR Direct Polymerase (Clontech, Mountain View, CA, USA)) improved the yield of cDNA (see Additional file , Figure S2c and Figure S5) and the reproducibility of the WTA replication (see Additional file , Figure S2d and Figure S5).

    Transformation Assay:

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Arabidopsis ecotype Col-0 (wild type) was transformed with Agrobacterium strain LBA4404 carrying either pMDC7::OsWRKY42 or pH 7FWG2::OsWRKY42 using the floral dip method [ ]. .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product.

    Translocation Assay:

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′). .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Transfection:

    Article Title: Control of alternative end joining by the chromatin remodeler p400 ATPase
    Article Snippet: After 48 h, siRNA transfection, cells were treated or not with 300 nM of 4-hydroxytamoxifen (4OHT) (Sigma; H7904) for 4 h. DNA was purified using QIAGEN DNeasy kit. .. Two microliters of digested or not samples (20 ng) were used as templates in 25 μl of qPCR reaction containing 12.5 μl TAKARA Mix PCR.

    Immunoprecipitation:

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: Paragraph title: pS6 immunoprecipitation and RNAseq ... 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The immunoprecipitated products were isolated with protein G agarose beads, and the bound chromatin was then collected by salt washing. .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C.

    Ligand Binding Assay:

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′). .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Infection:

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. The T2 progeny of three independent transgenic lines were used for functional studies.

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng). .. The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng).

    Generated:

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. Assays for callose deposition and bacterial infection were performed using inducible pMDC7::OsWRKY42 transgenic lines (XVE::OsWRKY42) that expressed high levels of OsWRKY42 .

    Article Title: Control of alternative end joining by the chromatin remodeler p400 ATPase
    Article Snippet: The level of resection generated at an AsiSI-induced DSB was measured by quantitative polymerase chain reaction (qPCR). .. Two microliters of digested or not samples (20 ng) were used as templates in 25 μl of qPCR reaction containing 12.5 μl TAKARA Mix PCR.

    Polymerase Chain Reaction:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: The nuclear ITS1 DNA was amplified by primers Betta_ITS1_F2 and Betta_ITS1_R2, Betta_ITS1_F3 and Betta_ITS1_R3, or Betta_ITS_F2 and Betta_ITS_R4 designed by Mahidol University (see ). .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. The PCR conditions were 98 °C for 2 min followed by 35 cycles of 98 °C for 10 s, annealing at 52 °C for 15 s and elongation at 68 °C for 1 min/kb.

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: Genomic DNA was isolated from cells/tissues using NucleoSpin Tissue kit (Macherey-Nagel) according to the manufacturer’s protocol. .. 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. Nested PCRs were performed under the same conditions using 5 μl of first PCR reaction.

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: The specificity and sensitivity of the PCR and potential interference by inhibitors in the feces were determined before assay application [ ]. .. Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. DNA fragments were amplified using the following optimized thermos-cycling conditions: 95 °C/5 min for denaturation; 35 cycles of 94 °C /30 s, 55 °C /30 s, 72 °C /40 s; and 72 °C /10 min extension.

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: The specificity and sensitivity of the PCR and potential interference by inhibitors in the feces were determined before assay application [ ]. .. Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × ExTaq Buffer, MgSO4 25 mM, 0.25 μl ExTaq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. DNA fragments were amplified using the following optimized thermos-cycling conditions: 95 °C/5 min for denaturation; 35 cycles of 94 °C /30 s, 55 °C /30 s, 72 °C /40 s; and 72 °C /10 min extension.

    Article Title: Whole-Genome Re-Alignment Facilitates Development of Specific Molecular Markers for Races 1 and 4 of Xanthomonas campestris pv. campestris, the Cause of Black Rot Disease in Brassica oleracea
    Article Snippet: The race 4 specific primers were: Xcc1_46R4 designed between 1843518 and 1843057 and Xcc1_46R4 primer designed between 1842956 to 1842379; these two primers were expected to amplify 462 and 578 base pairs, respectively ( ). .. Emerald PCR master mix (Takara, Shiga, Japan) was used in Polymerase Chain Reaction (PCR) for amplification of the target regions with respective markers. .. A 20.0 μL of PCR reaction mixture containing forward and reverse primers (1.0 μL each), Emerald PCR master mix (9.0 μL), ultra-pure water (8 μL) and 1.0 μL DNA was used for PCR amplification.

    Article Title: Plasmodium vivax aldolase-specific monoclonal antibodies and its application in clinical diagnosis of malaria infections in China
    Article Snippet: This opens the door to the production of P. vivax specific RDTs that target the aldolase antigen and can therefore used as an alternative to augment the already existing malaria RDTs. .. Restriction endonucleases, NdeI and SalI, and PrimeStar Hs mix, and PCR mix, DNA markers were purchased from TaKaRa Biotech Company (Dalian, China). pET-30a plasmid, competent E. coli cells (DH5α and BL21 (DE3)) and SP2/0-Ag14 myeloma cells were preserved in School of Bioscience and Bioengineering, South China University of Technology (China). .. RPMI 1640 medium, fetal bovine serum (FBS), kanamycin, penicillin-streptomycin, hypoxanthine and thymidine (HT), hypoxanthine-aminopterin-thymidine medium (HAT) and polyethylene glycol (PEG) were purchased from Gibco (California, U.S.A).

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 1.5 µl purified RNA was mixed with 5 µl reaction mix (1× PCR buffer (Roche), 1.5 mM MgCl2 , 50 µM dNTPs, 2 ng/µl poly–T primer (TATAGAATTCGCGGCCGCTCGCGATTTTTTTTTTTTTTTTTTTTTTTT), 0.04 U/µl RNase inhibitor (Qiagen), 0.4 U/µl recombinant RNasin (Promega)). .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min. .. The PCR product was purified by gel purification, and 50 ng of the purified product was used for library preparation with Nextera DNA Sample Prep kits (Illumina).

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Transgenic seedlings were selected on MS agar medium containing hygromycin (25 μg/ml). .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. The T2 progeny of three independent transgenic lines were used for functional studies.

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Age-matched syngeneic adult male mice (12–13-week-old; 22–27 g body weight) were used in this study. .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′). .. To achieve inactivation of Mdm2 and p53 in the adult mouse heart, we used the Cre- loxP recombination system of bacteriophage P1.

    Article Title: Mutation of kri1l causes definitive hematopoiesis failure via PERK-dependent excessive autophagy induction
    Article Snippet: RNA was reverse-transcribed using random hexamers and SuperScript III Reverse Transcriptase (Invitrogen). .. 2× PCR Mix (TaKaRa, Premix Ex Taq) containing SYBR Green I was used for the real-time quantitative PCR analysis with the Applied Biosystems 7900HT Fast Real-Time PCR System. .. The relative expression values were normalized against the internal control actin (QPCR primer sequences were listed in ).

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: PCR amplification of the corresponding genomic region surrounding each SNP locus was performed in a TaKaRa PCR thermal cycler (TaKaRa TP600, China). .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA. .. Cycling conditions included 1 cycle at 95°C for 5 min, 40 cycles at 95°C for 30 s, melting temperature for 45 s, 72°C for 1 min, and a final extension at 72°C for 10 min. To avoid contamination, negative controls were included in each PCR.

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: VNTR in WSSV ORF94 was analyzed with primer ORF94 (Table ). .. The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng). .. The thermal cycle program was as follows: 94 °C for 4 min, followed by 35 cycles of 94 °C for 30 s, 60 °C for 30 s, 72 °C for 90 s and a final extension step of 72 °C for 10 min. Amplification products were verified by 1.5% agarose gel electrophoresis.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: The V4 region of the 16S rRNA gene specific to the eubacterial communities was amplified from the extracted DNA samples . .. A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA). .. Using a Veriti 96-well thermocycler (Life Technologies, Carlsbad, CA), the amplification was performed at 98 °C for 3 min, followed by 25 cycles of 30 s at 98 °C, 30 s at 55 °C, and 45 s at 68 °C, with a final elongation stage of 4 min at 68 °C.

    Article Title: Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity
    Article Snippet: Using the combination of the three previously discussed improvement points, we successfully suppressed the synthesis of byproducts completely in a single-tube reaction (see Additional file , Figure S2e), without using topoisomerase V. Furthermore, we selected a robust PCR enzyme that was optimal for the single-tube reaction. .. The use of this DNA polymerase (MightyAmp DNA Polymerase (Takara Bio, Inc., Tokyo, Japan), also marketed as Terra PCR Direct Polymerase (Clontech, Mountain View, CA, USA)) improved the yield of cDNA (see Additional file , Figure S2c and Figure S5) and the reproducibility of the WTA replication (see Additional file , Figure S2d and Figure S5). .. In addition, by using this DNA polymerase (MightyAmp), the number of PCR cycles in WTA could be reduced.

    Article Title: Changing Patterns of Malaria Epidemiology between 2002 and 2010 in Western Kenya: The Fall and Rise of Malaria
    Article Snippet: From 2006 onward, only legs were used for PCR species identification. .. About one-third of the leg samples were analyzed by DNA extraction and amplification of the ribosomal DNA using the Terra-Direct PCR method (Clonetech, Mountain View, CA. .. The rest of the samples from 2006 to 2010 were analyzed by using Fast Tissue-to-PCR kit (Fermentas, Glen Burnie, MA) and individual specimen was identified by the standard PCR procedure .

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: HepG2 cell line of hepatoblastoma and E. coli strains DH5α were conserved in our laboratory. .. Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Article Title: Sensitive and powerful single-cell RNA sequencing using mcSCRB-seq
    Article Snippet: Purified cDNA was eluted in 17 µl and residual primers digested with Exonuclease I (Thermo Fisher) for 20 min at 37 °C. .. After heat inactivation for 10 min at 80 °C, 30 µl PCR master mix consisting of 1.25 U Terra direct polymerase (Clontech) 1.66 × Terra direct buffer and 0.33 µM SINGV6 primer (IDT) was added. .. PCR was cycled as given: 3 min at 98 °C for initial denaturation followed by 15 cycles of 15 s at 98 °C, 30 s at 65 °C, 4 min at 68 °C.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: All ChIP primers used in the standard PCR and quantitative real-time PCR experiment are listed in Table . .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    DNA Sequencing:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. After purification, the PCR products were ligated to pPrime cloning vector (5PRIME).

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

    Sequencing:

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. PCR using genomic DNA extracted from Tg26 rat blood samples was performed in one step using LTR F and nested Gag R primers.

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min. .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min.

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Transgenic seedlings were selected on MS agar medium containing hygromycin (25 μg/ml). .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. The T2 progeny of three independent transgenic lines were used for functional studies.

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: For analysis of REL2 cDNA sequence in rel2 mutant, total RNA was extracted from young leaves of WT and rel2 mutant respectively. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Binding Assay:

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: After antibody binding, the mixture was added to the washed beads and gently mixed, followed by incubation for 1 hr at 4°C with rotation. .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min.

    Imaging:

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA. .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    DNA Extraction:

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: Genomic DNA was collected and isolated from oral mucosa swabs using a DNA extraction kit (Tiangen, China). .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: Paragraph title: DNA Extraction and PCR Amplification ... A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA).

    Article Title: Changing Patterns of Malaria Epidemiology between 2002 and 2010 in Western Kenya: The Fall and Rise of Malaria
    Article Snippet: From 2006 onward, only legs were used for PCR species identification. .. About one-third of the leg samples were analyzed by DNA extraction and amplification of the ribosomal DNA using the Terra-Direct PCR method (Clonetech, Mountain View, CA. .. The rest of the samples from 2006 to 2010 were analyzed by using Fast Tissue-to-PCR kit (Fermentas, Glen Burnie, MA) and individual specimen was identified by the standard PCR procedure .

    RNA Sequencing Assay:

    Article Title: Quartz-Seq: a highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity
    Article Snippet: The use of this DNA polymerase (MightyAmp DNA Polymerase (Takara Bio, Inc., Tokyo, Japan), also marketed as Terra PCR Direct Polymerase (Clontech, Mountain View, CA, USA)) improved the yield of cDNA (see Additional file , Figure S2c and Figure S5) and the reproducibility of the WTA replication (see Additional file , Figure S2d and Figure S5). .. The use of this DNA polymerase (MightyAmp DNA Polymerase (Takara Bio, Inc., Tokyo, Japan), also marketed as Terra PCR Direct Polymerase (Clontech, Mountain View, CA, USA)) improved the yield of cDNA (see Additional file , Figure S2c and Figure S5) and the reproducibility of the WTA replication (see Additional file , Figure S2d and Figure S5).

    Mutagenesis:

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Paragraph title: Mdm2 and p53 conditional mutant mice ... Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Both RT-PCR and qRT-PCR were used to analyze the expression level of REL2 /Os10g0562700 in WT and rel2 mutant. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Isolation:

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: Genomic DNA was isolated from cells/tissues using NucleoSpin Tissue kit (Macherey-Nagel) according to the manufacturer’s protocol. .. 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel.

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Age-matched syngeneic adult male mice (12–13-week-old; 22–27 g body weight) were used in this study. .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′). .. To achieve inactivation of Mdm2 and p53 in the adult mouse heart, we used the Cre- loxP recombination system of bacteriophage P1.

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Paragraph title: RNA Isolation and Gene Expression Analysis ... The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: Genomic DNA was collected and isolated from oral mucosa swabs using a DNA extraction kit (Tiangen, China). .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: Fecal DNA (6 from baseline, 12 per treatment) were isolated from approximately 100 mg samples using a commercially available kit (Mag-Bind Soil DNA Kit; Omega Bio-tek, Inc., Norcross, GA) according to the manufacturer’s protocol with the following modifications: the raw sample was transferred to a 2 mL sterile Safe-Lock tube (Eppendorf, Hauppauge, NY) with 0.3 g of acid washed beads (Scientific Asset Management, Basking Ridge, NJ) and 300 µL of SLX-Mlus Buffer, followed by bead-beating at frequency of 20 for 10 min (QIAGEN Inc., Valencia, CA); after samples were mixed with RNase A and DS Buffer, samples were incubated in a 90 °C water bath for 8 min and occasionally vortexed; the samples were centrifuged at a speed of 5,000 × g for 10 min and the supernatant was transferred; finally, DNA was eluted with 130 µL of elution buffer at the last step. .. A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA).

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The immunoprecipitated products were isolated with protein G agarose beads, and the bound chromatin was then collected by salt washing. .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C.

    Purification:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water.

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. PCR using genomic DNA extracted from Tg26 rat blood samples was performed in one step using LTR F and nested Gag R primers.

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 1.5 µl purified RNA was mixed with 5 µl reaction mix (1× PCR buffer (Roche), 1.5 mM MgCl2 , 50 µM dNTPs, 2 ng/µl poly–T primer (TATAGAATTCGCGGCCGCTCGCGATTTTTTTTTTTTTTTTTTTTTTTT), 0.04 U/µl RNase inhibitor (Qiagen), 0.4 U/µl recombinant RNasin (Promega)). .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min.

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: HepG2 cell line of hepatoblastoma and E. coli strains DH5α were conserved in our laboratory. .. Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Article Title: Control of alternative end joining by the chromatin remodeler p400 ATPase
    Article Snippet: After 48 h, siRNA transfection, cells were treated or not with 300 nM of 4-hydroxytamoxifen (4OHT) (Sigma; H7904) for 4 h. DNA was purified using QIAGEN DNeasy kit. .. Two microliters of digested or not samples (20 ng) were used as templates in 25 μl of qPCR reaction containing 12.5 μl TAKARA Mix PCR.

    Article Title: Sensitive and powerful single-cell RNA sequencing using mcSCRB-seq
    Article Snippet: Purified cDNA was eluted in 17 µl and residual primers digested with Exonuclease I (Thermo Fisher) for 20 min at 37 °C. .. After heat inactivation for 10 min at 80 °C, 30 µl PCR master mix consisting of 1.25 U Terra direct polymerase (Clontech) 1.66 × Terra direct buffer and 0.33 µM SINGV6 primer (IDT) was added.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The eluted ChIP Elution Buffer was then digested with proteinase K and purified for PCR analysis. .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C.

    Reverse Transcription Polymerase Chain Reaction:

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice
    Article Snippet: Both RT-PCR and qRT-PCR were used to analyze the expression level of REL2 /Os10g0562700 in WT and rel2 mutant. .. The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Positron Emission Tomography:

    Article Title: Plasmodium vivax aldolase-specific monoclonal antibodies and its application in clinical diagnosis of malaria infections in China
    Article Snippet: This opens the door to the production of P. vivax specific RDTs that target the aldolase antigen and can therefore used as an alternative to augment the already existing malaria RDTs. .. Restriction endonucleases, NdeI and SalI, and PrimeStar Hs mix, and PCR mix, DNA markers were purchased from TaKaRa Biotech Company (Dalian, China). pET-30a plasmid, competent E. coli cells (DH5α and BL21 (DE3)) and SP2/0-Ag14 myeloma cells were preserved in School of Bioscience and Bioengineering, South China University of Technology (China). .. RPMI 1640 medium, fetal bovine serum (FBS), kanamycin, penicillin-streptomycin, hypoxanthine and thymidine (HT), hypoxanthine-aminopterin-thymidine medium (HAT) and polyethylene glycol (PEG) were purchased from Gibco (California, U.S.A).

    IA:

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: The V4 region of the 16S rRNA gene specific to the eubacterial communities was amplified from the extracted DNA samples . .. A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA). .. Using a Veriti 96-well thermocycler (Life Technologies, Carlsbad, CA), the amplification was performed at 98 °C for 3 min, followed by 25 cycles of 30 s at 98 °C, 30 s at 55 °C, and 45 s at 68 °C, with a final elongation stage of 4 min at 68 °C.

    Mouse Assay:

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: Paragraph title: Mdm2 and p53 conditional mutant mice ... Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Chromatin Immunoprecipitation:

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: All ChIP primers used in the standard PCR and quantitative real-time PCR experiment are listed in Table . .. The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    Plasmid Preparation:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water.

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. PCR using genomic DNA extracted from Tg26 rat blood samples was performed in one step using LTR F and nested Gag R primers.

    Article Title: Plasmodium vivax aldolase-specific monoclonal antibodies and its application in clinical diagnosis of malaria infections in China
    Article Snippet: This opens the door to the production of P. vivax specific RDTs that target the aldolase antigen and can therefore used as an alternative to augment the already existing malaria RDTs. .. Restriction endonucleases, NdeI and SalI, and PrimeStar Hs mix, and PCR mix, DNA markers were purchased from TaKaRa Biotech Company (Dalian, China). pET-30a plasmid, competent E. coli cells (DH5α and BL21 (DE3)) and SP2/0-Ag14 myeloma cells were preserved in School of Bioscience and Bioengineering, South China University of Technology (China). .. RPMI 1640 medium, fetal bovine serum (FBS), kanamycin, penicillin-streptomycin, hypoxanthine and thymidine (HT), hypoxanthine-aminopterin-thymidine medium (HAT) and polyethylene glycol (PEG) were purchased from Gibco (California, U.S.A).

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng). .. The crayfish samples and shrimp samples were investigated for WSSV infection by qPCR using the primer VP28 for all WSSV strains and WSSV-CN-Pc-166 for WSSV-CN-Pc (Table ).

    Article Title: Changing Patterns of Malaria Epidemiology between 2002 and 2010 in Western Kenya: The Fall and Rise of Malaria
    Article Snippet: Vector species were morphologically identified as either An. funestu s or An. gambiae s.l. .. About one-third of the leg samples were analyzed by DNA extraction and amplification of the ribosomal DNA using the Terra-Direct PCR method (Clonetech, Mountain View, CA.

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: HepG2 cell line of hepatoblastoma and E. coli strains DH5α were conserved in our laboratory. .. Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Software:

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. PCR using genomic DNA extracted from Tg26 rat blood samples was performed in one step using LTR F and nested Gag R primers.

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng). .. The recombinant plasmid standard was serial-diluted to a range of 10 to 108 copies per microliter.

    Electrophoresis:

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × ExTaq Buffer, MgSO4 25 mM, 0.25 μl ExTaq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Article Title: Whole-Genome Re-Alignment Facilitates Development of Specific Molecular Markers for Races 1 and 4 of Xanthomonas campestris pv. campestris, the Cause of Black Rot Disease in Brassica oleracea
    Article Snippet: Emerald PCR master mix (Takara, Shiga, Japan) was used in Polymerase Chain Reaction (PCR) for amplification of the target regions with respective markers. .. A 20.0 μL of PCR reaction mixture containing forward and reverse primers (1.0 μL each), Emerald PCR master mix (9.0 μL), ultra-pure water (8 μL) and 1.0 μL DNA was used for PCR amplification.

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA. .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    Multiplex Assay:

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Paragraph title: Multiplex PCR assay ... Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl.

    Recombinant:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water. .. After purification, the PCR products were ligated to pPrime cloning vector (5PRIME).

    Article Title: Variation in olfactory neuron repertoires is genetically controlled and environmentally modulated
    Article Snippet: 1.5 µl purified RNA was mixed with 5 µl reaction mix (1× PCR buffer (Roche), 1.5 mM MgCl2 , 50 µM dNTPs, 2 ng/µl poly–T primer (TATAGAATTCGCGGCCGCTCGCGATTTTTTTTTTTTTTTTTTTTTTTT), 0.04 U/µl RNase inhibitor (Qiagen), 0.4 U/µl recombinant RNasin (Promega)). .. 0.3 µl RT mix (170 U/µl Superscript II (ThermoFisher Scientific), 0.4 U/µl RNase inhibitor (Qiagen), 4 U/µl recombinant RNasin (Promega), 3 µg/µl T4 gene 32 protein (Roche)) was added to each tube and incubated at 37°C for 10 min then at 65°C for 10 min. 1 µl ExoI mix (2 U/µl ExoI (NEB), 1× PCR buffer (Roche), 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 15 min then 80°C for 15 min. 5 µl TdT mix (1.25 U/µl TdT (Roche), 0.1 U/µl RNase H (ThermoFisher Scientific), 1× PCR buffer (Roche), 3 mM dATP, 1.5 mM MgCl2 ) was added to each tube and incubated at 37°C for 20 min then 65°C for 10 min. 3.5 µl of the product was added to 27.5 µl PCR mix (1× LA Taq reaction buffer (TaKaRa, Japan), 0.25 mM dNTPs, 20 ng/µl poly–T primer, 0.05 U/µl LA Taq (TaKaRa)) and incubated at 95°C for 2 min, 37°C for 5 min, 72°C for 20 min, then 16 cycles of 95°C for 30 s, 67°C for 1 min, 72°C for 3 min with 6 s extension for each cycle, and then 72°C for 10 min. .. The PCR product was purified by gel purification, and 50 ng of the purified product was used for library preparation with Nextera DNA Sample Prep kits (Illumina).

    Article Title: Characterization and prevalence of a novel white spot syndrome viral genotype in naturally infected wild crayfish, Procambarus clarkii, in Shanghai, China
    Article Snippet: The 25 μl PCR reaction contained 12.5 μl of 2 × PCR mix (TaKaRa), 0.5 μl of each of the 10 mM primers, 9.5 μl double distilled water and 2 μl DNA template (approximately 200 ng). .. The crayfish samples and shrimp samples were investigated for WSSV infection by qPCR using the primer VP28 for all WSSV strains and WSSV-CN-Pc-166 for WSSV-CN-Pc (Table ).

    Agarose Gel Electrophoresis:

    Article Title: Excision of HIV-1 DNA by Gene Editing: A Proof-of-Concept In Vivo Study
    Article Snippet: Genomic DNA was isolated from cells/tissues using NucleoSpin Tissue kit (Macherey-Nagel) according to the manufacturer’s protocol. .. 25ng of genomic DNA was subjected to PCR using Terra PCR direct polymerase mix (Takara, Clontech) under the following PCR conditions: 98 °C for 3 minutes, 35 cycles (98 °C for 10s, 68 °C for 1.30 minutes), 68 °C for 5 minutes and resolved in 1% agarose gel. .. Nested PCRs were performed under the same conditions using 5 μl of first PCR reaction.

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × ExTaq Buffer, MgSO4 25 mM, 0.25 μl ExTaq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Article Title: Whole-Genome Re-Alignment Facilitates Development of Specific Molecular Markers for Races 1 and 4 of Xanthomonas campestris pv. campestris, the Cause of Black Rot Disease in Brassica oleracea
    Article Snippet: Emerald PCR master mix (Takara, Shiga, Japan) was used in Polymerase Chain Reaction (PCR) for amplification of the target regions with respective markers. .. A 20.0 μL of PCR reaction mixture containing forward and reverse primers (1.0 μL each), Emerald PCR master mix (9.0 μL), ultra-pure water (8 μL) and 1.0 μL DNA was used for PCR amplification.

    Article Title: Single nucleotide polymorphisms predisposing to asthma in children of Mauritian Indian and Chinese Han ethnicity
    Article Snippet: The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA. .. The reaction was performed in a final volume of 10 μL including 2.05 μL commercial PCR master mix (TaKaRa Ex Taq), 5 pmol of each primer, and 10 ng genomic DNA.

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA). .. Using a Veriti 96-well thermocycler (Life Technologies, Carlsbad, CA), the amplification was performed at 98 °C for 3 min, followed by 25 cycles of 30 s at 98 °C, 30 s at 55 °C, and 45 s at 68 °C, with a final elongation stage of 4 min at 68 °C.

    Article Title: Characterization of the promoter region of the bovine SIX1 gene: Roles of MyoD, PAX7, CREB and MyoG
    Article Snippet: The ChIP-PCR reaction mixtures had a total volume of 20 μL containing 10 μL of PCR Mix (TaKaRa), 0.4 μM each primers and 50 ng of template DNA and were performed under the following conditions: initial incubation of 5 min at 95 °C, followed by 34 cycles of 30 s at 95 °C, 30 s at 60 °C and 30 s at 72 °C. .. The input, immunoprecipitated and normal rabbit IgG products were added to each tube as a template.

    Transgenic Assay:

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection
    Article Snippet: Transgenic seedlings were selected on MS agar medium containing hygromycin (25 μg/ml). .. The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product. .. The T2 progeny of three independent transgenic lines were used for functional studies.

    Article Title: p53 and Mdm2 act synergistically to maintain cardiac homeostasis and mediate cardiomyocyte cell cycle arrest through a network of microRNAs
    Article Snippet: The mcm transgenic miceon a C57BL/6J background were obtained from Jackson (Bar Harbor, ME04609 USA; strain name: B6.FVB(129)-Tg(Myh6-cre/Esr1*)1Jmk/J; stock number: 005657). .. Genotyping was performed by PCR using alkaline hydrolysis of genomic DNA isolated from tail tips using the Terra PCR Direct Kit ( mcm , Mdm2f/f , p53f/f ; Clontech) or AccuStart II GelTrack PCR Supermix and the following oligonucleotides: mcm ( forward 5′-AGGTGGACCTGA TCATGGAG-3′; reverse 5′-ATACCGGAGATCATGCAAGC-3′) Mdm2f/f ( forward 5′-CTGTGTGAGCTGAGGGAGATGTG-3′; reverse 5′-CCTGGATTTAATCTGCAGCACTC-3′) p53f/f ( forward 5′-CACAAAAACAGGTTAAACCCAG-3′; reverse 5′-AGCACATAGGAGGC AGAGAC-3′).

    Concentration Assay:

    Article Title: Effects of dietary supplementation of formaldehyde and crystalline amino acids on gut microbial composition of nursery pigs
    Article Snippet: The V4 region of the 16S rRNA gene specific to the eubacterial communities was amplified from the extracted DNA samples . .. A PCR reaction (20 µL) consisted of 2 µL of template DNA, 0.5 µL each of both forward and reverse 16S rRNA V4 primers (final concentration 10.0 µM; Integrated DNA Technologies, Coralville, IA), 0.5 µL of Terra PCR Direct Polymerase Mix (Clontech Laboratories Inc., Mountain View, CA), 10 µL of 2× Terra PCR Direct Buffer (Clontech Laboratories Inc., Mountain View, CA) and 6.5 µL of nuclease-free water (Hoefer Inc., Holliston, MA). .. Using a Veriti 96-well thermocycler (Life Technologies, Carlsbad, CA), the amplification was performed at 98 °C for 3 min, followed by 25 cycles of 30 s at 98 °C, 30 s at 55 °C, and 45 s at 68 °C, with a final elongation stage of 4 min at 68 °C.

    Marker:

    Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
    Article Snippet: Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

    Staining:

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × Ex Taq Buffer, MgSO4 25 mM, 0.25 μl Ex Taq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Article Title: Estimating the prevalence of Echinococcus in domestic dogs in highly endemic for echinococcosis
    Article Snippet: Briefly, PCR amplification was performed in a 25 μl mixture containing 12.5 μl PCR mix (dNTPs 2.5 mM of each, 2.5 μl 10 × ExTaq Buffer, MgSO4 25 mM, 0.25 μl ExTaq DNA polymerase 5 U/μl) (TaKaRa, Dalian, Liaoning), 4 μl of primer mix, 2 μl DNA template, and ddH2 O added to 25 μl. .. For all the multiplex PCR assays, positive DNA (DNA templates of E. granulosus and E. multilocularis ), and negative (no-DNA) controls were included [ ].

    Fluorescence In Situ Hybridization:

    Article Title: Biodiversity of the Betta smaragdina (Teleostei: Perciformes) in the northeast region of Thailand as determined by mitochondrial COI and nuclear ITS1 gene sequences
    Article Snippet: 2.2.2 DNA was extracted from muscle tissue using approximately 5 mg of the fish specimen incubating with 180 μl of 50 mM NaOH at 95 °C for 10 min. .. Amplification was carried out with the total volume of 25 μl containing 2 μl of the crude extract, 0.6 μl of 10 μM of each COI primer or 0.75 μl of 10 μM of each ITS1 primer, 0.5 μl of Terra™ PCR Direct Polymerase Mix (1.25 U/μl, Clontech), 10 μl of 2 × Terra™ PCR Direct Buffer (with Mg2 + and dNTP), and sterile distilled water.

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    TaKaRa terra pcr direct polymerase kit
    Ectopic expression of OsWRKY42 suppresses salt and cellulase induced expression of JA biosynthesis and responsive genes in Arabidopsis <t>transgenic</t> lines. Expression of JA biosynthesis and response genes was measured in 35S::OsWRKY42 transgenic lines and wild type (Col-0) Arabidopsis plants after salt and cellulase treatment. a. For salt treatment, one-week old seedlings (wild type or 35S::OsWRKY42) were treated for 12 h with either MS or MS with 150 mM NaCl. b. For cellulase treatment, leaves of three weeks old Arabidopsis plants (wild type or 35S::OsWRKY42) were infiltrated with 2 U/ml of cellulase (Sigma) . Leaves were harvested after 12 h and processed for qPCR analysis. The graph represents the relative fold change using expression values from wild type or 35S::OsWRKY42 lines treated with water. c. Leaves of fourteen days old rice seedlings were syringe infiltrated with either LBA4404 or LBA4404/pH7FWG2::OsWRKY42. Twelve hours post treatment, leaves were harvested and processed for <t>qRT-PCR.</t> The relative fold change was calculated over control (leaves infiltrated only with LBA4404). AtUBQ5 and OsGAPDH were used as endogenous controls for rice and Arabidopsis, respectively. The average from three biological replicates is plotted on the graph. The error bar represents standard deviation. Data was analysed using the Student’s t -test for independent means. The asterisk above the bar indicates significant difference with p value
    Terra Pcr Direct Polymerase Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 34 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Ectopic expression of OsWRKY42 suppresses salt and cellulase induced expression of JA biosynthesis and responsive genes in Arabidopsis transgenic lines. Expression of JA biosynthesis and response genes was measured in 35S::OsWRKY42 transgenic lines and wild type (Col-0) Arabidopsis plants after salt and cellulase treatment. a. For salt treatment, one-week old seedlings (wild type or 35S::OsWRKY42) were treated for 12 h with either MS or MS with 150 mM NaCl. b. For cellulase treatment, leaves of three weeks old Arabidopsis plants (wild type or 35S::OsWRKY42) were infiltrated with 2 U/ml of cellulase (Sigma) . Leaves were harvested after 12 h and processed for qPCR analysis. The graph represents the relative fold change using expression values from wild type or 35S::OsWRKY42 lines treated with water. c. Leaves of fourteen days old rice seedlings were syringe infiltrated with either LBA4404 or LBA4404/pH7FWG2::OsWRKY42. Twelve hours post treatment, leaves were harvested and processed for qRT-PCR. The relative fold change was calculated over control (leaves infiltrated only with LBA4404). AtUBQ5 and OsGAPDH were used as endogenous controls for rice and Arabidopsis, respectively. The average from three biological replicates is plotted on the graph. The error bar represents standard deviation. Data was analysed using the Student’s t -test for independent means. The asterisk above the bar indicates significant difference with p value

    Journal: BMC Plant Biology

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection

    doi: 10.1186/s12870-018-1391-5

    Figure Lengend Snippet: Ectopic expression of OsWRKY42 suppresses salt and cellulase induced expression of JA biosynthesis and responsive genes in Arabidopsis transgenic lines. Expression of JA biosynthesis and response genes was measured in 35S::OsWRKY42 transgenic lines and wild type (Col-0) Arabidopsis plants after salt and cellulase treatment. a. For salt treatment, one-week old seedlings (wild type or 35S::OsWRKY42) were treated for 12 h with either MS or MS with 150 mM NaCl. b. For cellulase treatment, leaves of three weeks old Arabidopsis plants (wild type or 35S::OsWRKY42) were infiltrated with 2 U/ml of cellulase (Sigma) . Leaves were harvested after 12 h and processed for qPCR analysis. The graph represents the relative fold change using expression values from wild type or 35S::OsWRKY42 lines treated with water. c. Leaves of fourteen days old rice seedlings were syringe infiltrated with either LBA4404 or LBA4404/pH7FWG2::OsWRKY42. Twelve hours post treatment, leaves were harvested and processed for qRT-PCR. The relative fold change was calculated over control (leaves infiltrated only with LBA4404). AtUBQ5 and OsGAPDH were used as endogenous controls for rice and Arabidopsis, respectively. The average from three biological replicates is plotted on the graph. The error bar represents standard deviation. Data was analysed using the Student’s t -test for independent means. The asterisk above the bar indicates significant difference with p value

    Article Snippet: The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product.

    Techniques: Expressing, Transgenic Assay, Mass Spectrometry, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Standard Deviation

    Ectopic expression of OsWRKY42 enhances tolerance to salt stress in Arabidopsis transgenic lines. a. One-week-old TN-1 rice seedlings (n = 10) were dipped in each of the following: 150 mM NaCl solution, 20% ( w / v ) PEG-6000 and mock (water). Leaves were harvested 12 h post treatment and processed for qRT-PCR. Relative fold change was calculated over water treated samples and OsGAPDH was used as an internal control. b. For assaying salt tolerance, one-week-old Arabidopsis seedlings that are either constitutively expressing OsWRKY42 (35S::OsWRKY42) or wild type (Col-0) ( n = 20) were grown on MS agar medium with or without NaCl (150 mM, Merck). (c-d) The total fresh weight and number of root branches of individual seedlings was quantified on the fifteenth day post-treatment. e . For anthocyanin estimation, three weeks old Arabidopsis plants (wild type and 35S::OsWRKY42) were treated with either 150 mM NaCl or water. On the fifteenth day post treatment, leaves ( n = 3) were collected and anthocyanin was extracted in acidic methanol. Anthocyanin was estimated using a spectrophotometer and the anthocyanin content is expressed as absorbance per milligram of fresh weight (F.W). f . ROS estimation was done in detached leaves from adult plants. Leaf discs (n = 15) of either wild type or 35S::OsWRKY42 lines were placed on sterile filter papers soaked in MS solution either with or without NaCl (150 mM). 12 h post treatment, the leaves were stained with NBT. The O 2 − content was estimated spectrophotometrically and is expressed as change in absorbance per milligram of fresh weight. Leaves from three different plants were used as replicates. The data was analysed using one-way ANOVA and the Tukey-Karmer honestly significance difference test. The alphabets above the bar indicates significant difference with p value

    Journal: BMC Plant Biology

    Article Title: Overexpression of a cell wall damage induced transcription factor, OsWRKY42, leads to enhanced callose deposition and tolerance to salt stress but does not enhance tolerance to bacterial infection

    doi: 10.1186/s12870-018-1391-5

    Figure Lengend Snippet: Ectopic expression of OsWRKY42 enhances tolerance to salt stress in Arabidopsis transgenic lines. a. One-week-old TN-1 rice seedlings (n = 10) were dipped in each of the following: 150 mM NaCl solution, 20% ( w / v ) PEG-6000 and mock (water). Leaves were harvested 12 h post treatment and processed for qRT-PCR. Relative fold change was calculated over water treated samples and OsGAPDH was used as an internal control. b. For assaying salt tolerance, one-week-old Arabidopsis seedlings that are either constitutively expressing OsWRKY42 (35S::OsWRKY42) or wild type (Col-0) ( n = 20) were grown on MS agar medium with or without NaCl (150 mM, Merck). (c-d) The total fresh weight and number of root branches of individual seedlings was quantified on the fifteenth day post-treatment. e . For anthocyanin estimation, three weeks old Arabidopsis plants (wild type and 35S::OsWRKY42) were treated with either 150 mM NaCl or water. On the fifteenth day post treatment, leaves ( n = 3) were collected and anthocyanin was extracted in acidic methanol. Anthocyanin was estimated using a spectrophotometer and the anthocyanin content is expressed as absorbance per milligram of fresh weight (F.W). f . ROS estimation was done in detached leaves from adult plants. Leaf discs (n = 15) of either wild type or 35S::OsWRKY42 lines were placed on sterile filter papers soaked in MS solution either with or without NaCl (150 mM). 12 h post treatment, the leaves were stained with NBT. The O 2 − content was estimated spectrophotometrically and is expressed as change in absorbance per milligram of fresh weight. Leaves from three different plants were used as replicates. The data was analysed using one-way ANOVA and the Tukey-Karmer honestly significance difference test. The alphabets above the bar indicates significant difference with p value

    Article Snippet: The transgenic nature of the T1 plants was further confirmed by direct PCR (Terra PCR direct polymerase kit, Clontech using leaf tissue and sequencing of the amplified product.

    Techniques: Expressing, Transgenic Assay, Quantitative RT-PCR, Mass Spectrometry, Spectrophotometry, Staining

    Antiviral function of HM- γ -PGA in murine macrophage cell line. (a) GFP expression images of medium only, 1 mg/mL HM- γ -PGA, and 1000 units/mL recombinant mouse IFN- β treated cells 12 h before NDV-GFP infection. Images taken 12 hpi (200x magnification). (b1) Viral mRNA expression level of Matrix gene of NDV-gfp over time in each treatment group was confirmed by specific RT-PCR primers which are shown in Table 1 . Equal amounts of PCR products were run on 1.5% ethidium bromide impregnated agarose gels and visualized using GelDoc Imaging System. All samples were normalized using their respective GAPDH gene expression. (b2) Relative band intensity (RBI) of the Matrix gene mRNA expression of (b1). RBI was determined (Gene/GAPDH) using GelDoc Imaging System Band Quantification Software. Error bars indicate the range of values obtained from two independent experiments. (c) GFP expression level of media treated, 1 mg/mL HM- γ -PGA, and IFN- β treated cells 12 h before NDV-GFP (La Sota strain) infection. GFP expression was measured 24 hpi using Glomax multidetection system. (d) Cell viability was determined by trypan blue exclusion test at 30 hpi. The results are presented as a percentage of the control (cells without treatment). Error bars on Figures (c) and (d) indicate the range of values obtained from triplicate counting ( ** P

    Journal: Advances in Virology

    Article Title: The Antiviral Effect of High-Molecular Weight Poly-Gamma-Glutamate against Newcastle Disease Virus on Murine Macrophage Cells

    doi: 10.1155/2014/301386

    Figure Lengend Snippet: Antiviral function of HM- γ -PGA in murine macrophage cell line. (a) GFP expression images of medium only, 1 mg/mL HM- γ -PGA, and 1000 units/mL recombinant mouse IFN- β treated cells 12 h before NDV-GFP infection. Images taken 12 hpi (200x magnification). (b1) Viral mRNA expression level of Matrix gene of NDV-gfp over time in each treatment group was confirmed by specific RT-PCR primers which are shown in Table 1 . Equal amounts of PCR products were run on 1.5% ethidium bromide impregnated agarose gels and visualized using GelDoc Imaging System. All samples were normalized using their respective GAPDH gene expression. (b2) Relative band intensity (RBI) of the Matrix gene mRNA expression of (b1). RBI was determined (Gene/GAPDH) using GelDoc Imaging System Band Quantification Software. Error bars indicate the range of values obtained from two independent experiments. (c) GFP expression level of media treated, 1 mg/mL HM- γ -PGA, and IFN- β treated cells 12 h before NDV-GFP (La Sota strain) infection. GFP expression was measured 24 hpi using Glomax multidetection system. (d) Cell viability was determined by trypan blue exclusion test at 30 hpi. The results are presented as a percentage of the control (cells without treatment). Error bars on Figures (c) and (d) indicate the range of values obtained from triplicate counting ( ** P

    Article Snippet: The polymerase chain reactions (PCR) using specific primers for Matrix gene of APMV-1 [ ] and murine glyceraldehyde 3-phosphate dehydrogenase (GAPDH) housekeeping gene (internal control) [ ] were carried out using emerald PCR master mix (Takara, USA) with 1 pM of each primer set ( ).

    Techniques: Expressing, Recombinant, Infection, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Imaging, Software

    Induction of antiviral genes and proinflammatory cytokines by HM- γ -PGA in RAW 264.7 cells. (a) Cells were treated with medium only, HM- γ -PGA (1 mg/mL), and 100 ng/mL of LPS. The time-dependent changes in mRNA expression after treatment were confirmed by PCR. All samples were normalized using GAPDH wherein equal amounts PCR products were run on 1.5% ethidium bromide impregnated agarose gels and visualized using GelDoc Imaging System. (b) RBI (Gene/GAPDH) of the IFN- β , IRF-3, IRF-7, and IFN-related genes of Figure (a). RBI was determined using GelDoc Imaging System Band Quantification Software. Error bars indicate the range of values obtained from two independent experiments.

    Journal: Advances in Virology

    Article Title: The Antiviral Effect of High-Molecular Weight Poly-Gamma-Glutamate against Newcastle Disease Virus on Murine Macrophage Cells

    doi: 10.1155/2014/301386

    Figure Lengend Snippet: Induction of antiviral genes and proinflammatory cytokines by HM- γ -PGA in RAW 264.7 cells. (a) Cells were treated with medium only, HM- γ -PGA (1 mg/mL), and 100 ng/mL of LPS. The time-dependent changes in mRNA expression after treatment were confirmed by PCR. All samples were normalized using GAPDH wherein equal amounts PCR products were run on 1.5% ethidium bromide impregnated agarose gels and visualized using GelDoc Imaging System. (b) RBI (Gene/GAPDH) of the IFN- β , IRF-3, IRF-7, and IFN-related genes of Figure (a). RBI was determined using GelDoc Imaging System Band Quantification Software. Error bars indicate the range of values obtained from two independent experiments.

    Article Snippet: The polymerase chain reactions (PCR) using specific primers for Matrix gene of APMV-1 [ ] and murine glyceraldehyde 3-phosphate dehydrogenase (GAPDH) housekeeping gene (internal control) [ ] were carried out using emerald PCR master mix (Takara, USA) with 1 pM of each primer set ( ).

    Techniques: Expressing, Polymerase Chain Reaction, Imaging, Software

    qRT-PCR analysis of the expression pattern of REL2 . Total RNA was extracted from various tissues from callus and WT plants. Rice Actin1 gene was used as a control. The expression of REL2 in roots at heading stage was set to 1. The 1 st leaf blades or 2 nd leaf blade mean the position of leaf blades or leaf sheaths. From up to down of a rice plant, the first leaf blade or leaf sheath is the 1 st leaf blade or leaf sheath and the second leaf blade or leaf sheath is the 2 nd leaf blade or leaf sheath. The values are the mean ± SEM with three biological replicates

    Journal: Rice

    Article Title: REL2, A Gene Encoding An Unknown Function Protein which Contains DUF630 and DUF632 Domains Controls Leaf Rolling in Rice

    doi: 10.1186/s12284-016-0105-6

    Figure Lengend Snippet: qRT-PCR analysis of the expression pattern of REL2 . Total RNA was extracted from various tissues from callus and WT plants. Rice Actin1 gene was used as a control. The expression of REL2 in roots at heading stage was set to 1. The 1 st leaf blades or 2 nd leaf blade mean the position of leaf blades or leaf sheaths. From up to down of a rice plant, the first leaf blade or leaf sheath is the 1 st leaf blade or leaf sheath and the second leaf blade or leaf sheath is the 2 nd leaf blade or leaf sheath. The values are the mean ± SEM with three biological replicates

    Article Snippet: The primers were listed in additional file : Table S3. qRT-PCR was carried out using the real-time PCR master mix (SYBR green, TaKaRa) with the CFX96 machine (Bio-Rad) following the manufacturer’s instructions.

    Techniques: Quantitative RT-PCR, Expressing

    PCR confirming the natural transformation in E. anophelis endophthalmitis. Lane 1: 1 kb DNA ladder. Lane 2: Positive control (pCMV-6 BDNF plasmid used as template). Expected band size of BDNF gene is 750 bp. Lane 3–5: PCR products from plasmid isolated from E. anophelis endophthalmitis after natural transformation. Lane 6–7: Negative controls.

    Journal: Scientific Reports

    Article Title: Comparative genomic analysis of a naturally competent Elizabethkingia anophelis isolated from an eye infection

    doi: 10.1038/s41598-018-26874-8

    Figure Lengend Snippet: PCR confirming the natural transformation in E. anophelis endophthalmitis. Lane 1: 1 kb DNA ladder. Lane 2: Positive control (pCMV-6 BDNF plasmid used as template). Expected band size of BDNF gene is 750 bp. Lane 3–5: PCR products from plasmid isolated from E. anophelis endophthalmitis after natural transformation. Lane 6–7: Negative controls.

    Article Snippet: PCR was performed for the presence of BDNF gene in the plasmids isolated from the transformaned colonies using specific primers (forward Primer: 5′-GGATCCATGACCATCCTTTTCCTTACTATGG-3′; reverse Primer: 5′-AAGCTTCTATCTTCCCCTTTTAATGGTCAGT-3′) in a final volume of 20 μl containing 10 μl of PCR Master Mix (Takara) which includes dNTPs, MgCl2, Taq DNA polymerase and PCR buffer), 0.5 μM of forward and reverse primers, template DNA (2 μl) and nuclease free Water (4 μl).

    Techniques: Polymerase Chain Reaction, Transformation Assay, Positive Control, Plasmid Preparation, Isolation