Journal: Methods and Protocols

Article Title: Comparison of Commercially Available Thermostable DNA Polymerases with Reverse Transcriptase Activity in Coupled Reverse Transcription Polymerase Chain Reaction Assays

doi: 10.3390/mps8010011

Figure Lengend Snippet: End-point RT-PCR assays with RevTaq, OmniTaq 2, and ReverHotTaq DNA polymerases and OneTaq One-Step Enzyme Mix. The indicated DNA polymerases and OneTaq enzyme mixture were used for the RT-PCR amplification of 105 bp, 203 bp, and 317 bp fragments (indicated by arrows) of the human beta-2-microglobulin mRNA sequence. The reaction mixtures contained a human total RNA as a template: 10 ng—lane 1; 1 ng—lane 2; 100 pg—lane 3; 10 pg—lane 4; no template control (NTC)—lane 5.

Article Snippet: The following materials were obtained from these sources: 100X OmniTaq2 DNA polymerase and 10X Taq Mutant Reaction Buffer from DNA Polymerase Technology, Inc. (St. Louis, MO, USA); 50X RevTaq DNA polymerase and 5X Volcano Reaction Buffer from myPOLS Biotec GmbH (Konstanz, Germany); 50X ReverHotTaq polymerase and 5X Reaction Buffer from Bioron GmbH (Römerberg, Germany); OneTaq One-Step RT-PCR Kit from New England BioLabs, Inc. (Ipswich, MA, USA); 5X OneTube RT-PCR-Mix from Evrogen (Moscow, Russia); and dNTPs from Bioline Limited (London, UK).

Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification, Sequencing, Control

Journal: Methods and Protocols

Article Title: Comparison of Commercially Available Thermostable DNA Polymerases with Reverse Transcriptase Activity in Coupled Reverse Transcription Polymerase Chain Reaction Assays

doi: 10.3390/mps8010011

Figure Lengend Snippet: Real-time RT-qPCR with EvaGreen intercalating dye and melting curves of amplicons. The RT-qPCR assays of human GAPDH mRNA in the total RNA were carried out with the indicated enzymes: ReverHotTaq ( A ), RevTaq ( B ), and OmniTaq2 ( C ) DNA polymerases and OneTube RT-PCRMix ( D ). The reaction mixtures contained 1 ng (brown curves), 0.1 ng (beige curves), or 10 pg (green curves) of human total RNA per reaction, or no template NTC (gray curves). Melting curves demonstrate the yields of specific (melting peaks over 80 °C) and non-specific (melting peaks below 80 °C) amplicons.

Article Snippet: The following materials were obtained from these sources: 100X OmniTaq2 DNA polymerase and 10X Taq Mutant Reaction Buffer from DNA Polymerase Technology, Inc. (St. Louis, MO, USA); 50X RevTaq DNA polymerase and 5X Volcano Reaction Buffer from myPOLS Biotec GmbH (Konstanz, Germany); 50X ReverHotTaq polymerase and 5X Reaction Buffer from Bioron GmbH (Römerberg, Germany); OneTaq One-Step RT-PCR Kit from New England BioLabs, Inc. (Ipswich, MA, USA); 5X OneTube RT-PCR-Mix from Evrogen (Moscow, Russia); and dNTPs from Bioline Limited (London, UK).

Techniques: Quantitative RT-PCR

Journal: Methods and Protocols

Article Title: Comparison of Commercially Available Thermostable DNA Polymerases with Reverse Transcriptase Activity in Coupled Reverse Transcription Polymerase Chain Reaction Assays

doi: 10.3390/mps8010011

Figure Lengend Snippet: Comparison of the indicated enzymes in RT-qPCR with EvaGreen intercalating dye (A) and with TaqMan probe (B). The mean quantification cycles (Cq) ± standard deviation of three replicates for the indicated template total RNA amounts (NTC, no template control) are provided. ∆Cq is the difference between the Cq of the RT-qPCR with the indicated polymerase and control OneTube RT-PCR-Mix for each template dilution. (A) The real-time amplifications of GAPDH mRNA fragment with EvaGreen intercalating dye were carried out using 10-fold dilutions of the human total RNA from 1 ng to 0.01 ng as a template, or no template control (NTC). (B) The TaqMan prob RT-qPCR assays of SARS-CoV-2 viral RNA were carried out with 10, 50, and 250 times diluted RNA isolated from SARS-CoV-2-positive nasopharyngeal swabs as a template or no template control (NTC).

Article Snippet: The following materials were obtained from these sources: 100X OmniTaq2 DNA polymerase and 10X Taq Mutant Reaction Buffer from DNA Polymerase Technology, Inc. (St. Louis, MO, USA); 50X RevTaq DNA polymerase and 5X Volcano Reaction Buffer from myPOLS Biotec GmbH (Konstanz, Germany); 50X ReverHotTaq polymerase and 5X Reaction Buffer from Bioron GmbH (Römerberg, Germany); OneTaq One-Step RT-PCR Kit from New England BioLabs, Inc. (Ipswich, MA, USA); 5X OneTube RT-PCR-Mix from Evrogen (Moscow, Russia); and dNTPs from Bioline Limited (London, UK).

Techniques: Comparison, Standard Deviation, Control, Isolation

Journal: Methods and Protocols

Article Title: Comparison of Commercially Available Thermostable DNA Polymerases with Reverse Transcriptase Activity in Coupled Reverse Transcription Polymerase Chain Reaction Assays

doi: 10.3390/mps8010011

Figure Lengend Snippet: Real-time RT-qPCR with TaqMan probe. The RT-qPCR assays of SARS-CoV-2 viral RNA were carried out with the indicated enzymes: ReverHotTaq ( A ), RevTaq ( C ), and OmniTaq2 ( D ) DNA polymerases and OneTube RT-PCRMix ( B ). The reaction mixtures contained 10 times diluted (brown curves), 50 times diluted (beige curves), or 250 times diluted (green curves) sample of RNA isolated from SARS-CoV-2-positive nasopharyngeal swabs as a template or no template NTC (gray curves).

Article Snippet: The following materials were obtained from these sources: 100X OmniTaq2 DNA polymerase and 10X Taq Mutant Reaction Buffer from DNA Polymerase Technology, Inc. (St. Louis, MO, USA); 50X RevTaq DNA polymerase and 5X Volcano Reaction Buffer from myPOLS Biotec GmbH (Konstanz, Germany); 50X ReverHotTaq polymerase and 5X Reaction Buffer from Bioron GmbH (Römerberg, Germany); OneTaq One-Step RT-PCR Kit from New England BioLabs, Inc. (Ipswich, MA, USA); 5X OneTube RT-PCR-Mix from Evrogen (Moscow, Russia); and dNTPs from Bioline Limited (London, UK).

Techniques: Quantitative RT-PCR, Isolation

Journal: Experimental and Therapeutic Medicine

Article Title: Expression and significance of NELIN and SM22α in varicose vein tissue

doi: 10.3892/etm.2015.2170

Figure Lengend Snippet: Primer sequences used for reverse transcription polymerase chain reaction analysis of NELIN.

Article Snippet: TransZol Up and two-step reverse transcription polymerase chain reaction (RT-PCR) kits were purchased from Beijing TransGen Biotech Co., Ltd. (Beijing, China). β-actin upstream and downstream primers were synthesized by Takara Bio, Inc. (Shiga, Japan).

Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification

Journal: Experimental and Therapeutic Medicine

Article Title: Expression and significance of NELIN and SM22α in varicose vein tissue

doi: 10.3892/etm.2015.2170

Figure Lengend Snippet: Primer sequences used for reverse transcription polymerase chain reaction analysis of SM22α.

Article Snippet: TransZol Up and two-step reverse transcription polymerase chain reaction (RT-PCR) kits were purchased from Beijing TransGen Biotech Co., Ltd. (Beijing, China). β-actin upstream and downstream primers were synthesized by Takara Bio, Inc. (Shiga, Japan).

Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification

Journal: Experimental and Therapeutic Medicine

Article Title: Expression and significance of NELIN and SM22α in varicose vein tissue

doi: 10.3892/etm.2015.2170

Figure Lengend Snippet: Reverse transcription polymerase chain reaction analysis revealed weak mRNA expression of NELIN in the vascular smooth muscle cells (VSMCs) of the experimental group, but strong expression in the control group. (A) Expected length of the PCR products was 611 bp (NELIN) and 312 bp (β-actin). Lanes: N, control group; X, DNA marker DL 2000; and P, experimental group. (B) Downregulation of NELIN mRNA expression in the VSMCs from the varicose vein tissue. Data are presented as the mean ± standard deviation.

Article Snippet: TransZol Up and two-step reverse transcription polymerase chain reaction (RT-PCR) kits were purchased from Beijing TransGen Biotech Co., Ltd. (Beijing, China). β-actin upstream and downstream primers were synthesized by Takara Bio, Inc. (Shiga, Japan).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Marker, Standard Deviation

Journal: Experimental and Therapeutic Medicine

Article Title: Expression and significance of NELIN and SM22α in varicose vein tissue

doi: 10.3892/etm.2015.2170

Figure Lengend Snippet: Reverse transcription polymerase chain reaction analysis revealed weak mRNA expression of SM22α in the vascular smooth muscle cells (VSMCs) of the experimental group, but strong expression in the control group. (A) Expected length of the PCR products was 232 bp (SM22α) and 472 bp (GADPH). Lanes: N, control group; X, DNA marker DL 2000; and P, experimental group. (B) Downregulation of SM22α mRNA expression in the VSMCs from the varicose vein tissue. Data are presented as the mean ± standard deviation.

Article Snippet: TransZol Up and two-step reverse transcription polymerase chain reaction (RT-PCR) kits were purchased from Beijing TransGen Biotech Co., Ltd. (Beijing, China). β-actin upstream and downstream primers were synthesized by Takara Bio, Inc. (Shiga, Japan).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Marker, Standard Deviation