Sensitive Aptamer SERS and RRS Assays for Trace Oxytetracycline Based on the Catalytic Amplification of CuNCs

A new method for the determination of oxytetracycline (OTC) has been established by coupling the catalytic amplification reaction of copper nanoclusters (CuNCs) with the aptamer reaction. CuNCs prepared by a wet chemical method have the catalytic activity for the formation of gold nanoparticles (AuNPs) resulting from a HAuCl4-ethanol (En) reaction. The experimental results showed that OTC aptamer (Apt) can be adsorbed on the surface of CuNCs in a non-specific way, thus inhibiting its catalytic activity. When OTC was added to the solution, the OTC-Apt complex was generated by a specific reaction, which made the CuNCs desorb and restore their catalytic activity. With the increase of OTC, the recovery of the catalytic activity of CuNCs is strengthened, the reaction speed is accelerated, and the number of AuNPs is increased. The generated AuNPs exhibited surface enhanced Raman scattering (SERS) signals at 1615 cm−1 in the presence of Vitoria blue 4R (VB4R) molecular probes, and a resonance Rayleigh scattering (RRS) peak at 586 nm. There is a good linear relationship between the intensities of SERS, or RRS, and OTC concentration at the range of 37.5–300 ng/L or 37.5–225 ng/L, respectively. A new SERS and RRS assay for the determination of trace OTC based on the regulation of CuNCs catalysis was established.

NEBNext® Varskip Short ARTIC SARS-CoV-2 RT-PCR Module E7626 v1

This protocol details methods for the NEBNext® ARTIC SARS-CoV-2 RT-PCR Module, NEB #E7626S/L 24/96 reactions. cDNA Synthesis and Targeted cDNA Amplification with NEBNext VarSkip Short Primer Mixes: This protocol follows an alternate variant-tolerant approach for targeting SARS-CoV-2 by utilizing NEBNext VarSkip Short SARS-CoV-2 Primer Mixes. The NEBNext VarSkip Short SARS-CoV-2 Primer mixes cannot be added to the same cDNA amplification reaction as the NEBNext ARTIC SARS-CoV-2 Primer Mixes. If downstream applications include sequencing, performing RNA input normalization prior to cDNA synthesis and targeted amplification promotes more even distribution of reads across sequencing libraries. For other NEBNext® ARTIC SARS-CoV-2 protocols, please see the NEBNext ARTIC Protocols Collection. To obtain instructions for using NEBNext ARTIC SARS-CoV-2 Primer Mix and the NEBNext® ARTIC SARS-CoV-2 RT-PCR Module please see the NEBNext ARTIC SARS CoV2 RT PCR Module Manual.

NEBNext® Varskip Short ARTIC SARS-CoV-2 FS Library Prep Kit (Illumina®) E7658 Express Protocol with One Clean-up Step v1

This protocol details methods for the NEBNext® ARTIC SARS-CoV-2 FS Library Prep Kit (Illumina®), NEB #E7658S/L 24/96 reactions. This protocol follows an alternate variant-tolerant approach for targeting SARS-CoV-2 by utilizing NEBNext VarSkip Short SARS-CoV-2 Primer Mixes. The NEBNext VarSkip Short SARS-CoV-2 Primer mixes cannot be added to the same cDNA amplification reaction as the NEBNext ARTIC SARS-CoV-2 Primer Mixes. This protocol does not include a cleanup step for each sample after cDNA synthesis and after adaptor ligation. Performing RNA input normalization prior to cDNA synthesis and targeted amplification and/or normalizing final libraries prior to sequencing promotes more even distribution of reads across libraries. Skipping RNA input normalization, final library normalization, and cleanups reduces hands on time but may require deeper sequencing depth to reach sufficient coverage of each sample. For other NEBNext® ARTIC SARS-CoV-2 protocols, please see the NEBNext ARTIC Protocols Collection.

Ultrarapid detection of SARS-CoV-2 RNA using a reverse transcription–free exponential amplification reaction, RTF-EXPAR

A rapid isothermal method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for COVID-19, is reported. The procedure uses an unprecedented reverse transcription–free (RTF) approach for converting genomic RNA into DNA. This involves the formation of an RNA/DNA heteroduplex whose selective cleavage generates a short DNA trigger strand, which is then rapidly amplified using the exponential amplification reaction (EXPAR). Deploying the RNA-to-DNA conversion and amplification stages of the RTF-EXPAR assay in a single step results in the detection, via a fluorescence read-out, of single figure copy numbers per microliter of SARS-CoV-2 RNA in under 10 min. In direct three-way comparison studies, the assay has been found to be faster than both RT-qPCR and reverse transcription loop-mediated isothermal amplification (RT-LAMP), while being just as sensitive. The assay protocol involves the use of standard laboratory equipment and is readily adaptable for the detection of other RNA-based pathogens.