rna polymerase (New England Biolabs)


Structured Review

Rna Polymerase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rna polymerase/product/New England Biolabs
Average 93 stars, based on 1 article reviews
Price from $9.99 to $1999.99
Images
1) Product Images from "SCOPE enables type III CRISPR-Cas diagnostics using flexible targeting and stringent CARF ribonuclease activation"
Article Title: SCOPE enables type III CRISPR-Cas diagnostics using flexible targeting and stringent CARF ribonuclease activation
Journal: Nature Communications
doi: 10.1038/s41467-021-25337-5

Figure Legend Snippet: A novel type III CRISPR-Cas tool for the sensitive detection of nucleic acids. a Denaturing PAGE resulting from activity assays performed with the CARF protein TTHB144 and a 5′ Cy5-labeled reporter RNA. Activation of TTHB144 due to cOAs produced by the reconstituted TtCmr-46 complex was monitored by offering fully complementary (T) target RNAs, or target RNA with mismatches in segments one (S1), four (S4) or by a single mismatch in the CAR (C1). A fully non-target RNA (NT) was used as a control. Results of the TTHB144 cleavage assay is representative of results obtained from three replicates (Supplementary Fig. S6 ). b Limit of detection (LOD) assay using reconstituted Cmr-46 on a synthetic SARS-CoV-2 E-gene and a fluorophore-quencher reporter RNA masking construct measured over time. A non-target RNA (NT) was used as a control. Data was obtained from three replicates. c Schematic overview of the 2-step reaction setup consisting out of (1) a (RT)-LAMP based pre-amplification step and (2) a T7-based in vitro transcription and type III CRISPR detection step. ‘F-Q’ represents the fluorophore-quencher reporter RNA. d Limit of detection assay using reconstituted Cmr-46 in the 2-step setup (depicted in panel c ), with a SARS-CoV-2 synthetic full RNA genome as target. Data was obtained from three replicates. e Detection of SARS-CoV-2 in human swab samples. Ct-values of qPCR analysis ( Orf1ab gene) of 81 samples are depicted on the X-axis with the true negative samples displayed as not determined (ND). See Supplementary Fig. S5 and Supplementary Table S2 for Ct-values of qPCR analysis of SARS-CoV-2 samples and respective Scope tool score. The data on the qPCR and SCOPE analysis was obtained from one replicate measurement. f One-pot LAMP-CRISPR limit of detection assay, using reconstituted Cmr-46, on a synthetic SARS-CoV-2 E-gene. Data was obtained from two replicates. All error bars in this figure represent the standard deviation of the mean. ‘Arb. units’ stands for arbitrary units. Source data are provided as a Source data file.
Techniques Used: CRISPR, Polyacrylamide Gel Electrophoresis, Activity Assay, Labeling, Activation Assay, Produced, Cleavage Assay, Construct, Amplification, In Vitro, Detection Assay, Real-time Polymerase Chain Reaction, Standard Deviation
2) Product Images from "Intrinsic Signal Amplification by Type-III CRISPR-Cas Systems Provides a Sequence-Specific Viral Diagnostic"
Article Title: Intrinsic Signal Amplification by Type-III CRISPR-Cas Systems Provides a Sequence-Specific Viral Diagnostic
Journal: medRxiv
doi: 10.1101/2020.10.14.20212670

Figure Legend Snippet: CRISPR-Csm-based detection of SARS-CoV-2 in clinical nasopharyngeal swab samples. ( A ) Schematic comparing RT-qPCR to RT-LAMP-CRISPR-Csm based detection. In RT-qPCR reaction , reverse transcribed template is amplified in PCR reaction that cycles between different temperatures. As Taq DNA polymerase (DNA pol.) synthesizes nascent DNA strand it degrades the probe annealed to amplified template and releases the fluorophore (green star) from the quencher (black hexagon). In CRISPR-Csm detection , the viral RNA is reverse transcribed, and resulting DNA is amplified in an RT-LAMP reaction to produce transcription templates for T7 RNA polymerase, in one pot. An aliquot of the RT-LAMP-T7 reaction (40 min) is then mixed with CRISPR-Csm reaction (20 min). ( B ) Nasopharyngeal swabs from 24 individuals were tested with RT-qPCR and RT-LAMP-T7 CRISPR-Csm. Top panel shows threshold cycle (Ct) values for RT-qPCR. Swabs with Ct values below 40 for both N1 and N2 CDC diagnostic primers are considered positive for SARS-CoV-2 RNA (nd – signal is not detected over during 45 cycles). Vertical dashed line highlights the Ct value corresponding to a concentration of 10 6 SARS-CoV-2 RNA copies/mL ( Supplemental Fig. 4 ). Bottom panel shows CRISPR-Csm based detection in the same swab samples targeting nucleocapsid (N) gene, as shown in (A). Data is shown as fold change in fluorescence compared to a no-template control reaction. Technical repeats are shown in triplicate. * - swab sample test result was inconclusive both for CDC assay and CRISPR-Csm detection.
Techniques Used: CRISPR, Quantitative RT-PCR, Amplification, Polymerase Chain Reaction, Diagnostic Assay, Concentration Assay, Fluorescence, CDC Assay
3) Product Images from "Inexpensive and colorimetric RNA detection by E. coli cell-free protein synthesis platform at room temperature"
Article Title: Inexpensive and colorimetric RNA detection by E. coli cell-free protein synthesis platform at room temperature
Journal: medRxiv
doi: 10.1101/2021.11.29.21267025

Figure Legend Snippet: Optimization and screening of isothermal RNA amplification platforms. (A) Non-specific products were detected with NASBA reaction, in parallel with previous reports 30 (B) Homemade NASBA reaction additive screening. (C) Influence of random RNA duplex on T7 RNA polymerase efficiency. Please note that more pronounced RNA bands were observed with the additions of the random duplex, in comparison to (A) and (B). (D) Influence of different reaction conditions and temperature cycling. (D) Room temperature amplifications by NASBA and RT-RPA. (Note that prior to loading onto gel, RT-RPA reactions –but not NASBA– were pre-incubated at 95 °C for 3 min and immediately placed on ice, in order to dissociate the SSB from the amplicons.) We also note that under reported reaction conditions, our initial attempts for RT-RCA did not yield amplification.
Techniques Used: Amplification, Recombinase Polymerase Amplification, Incubation