Template-switching polymerase chain reaction (TS-PCR) is a method of

reverse transcription A reverse transcriptase (RT) is an enzyme used to generate complementary DNA (cDNA) from an RNA template, a process termed reverse transcription. Reverse transcriptases are used by viruses such as HIV and hepatitis B to replicate their genomes, ...

and polymerase chain reaction (PCR) amplification that relies on a natural

PCR primer PCR or pcr may refer to: Science * Phosphocreatine, a phosphorylated creatine molecule * Principal component regression, a statistical technique Medicine * Polymerase chain reaction ** COVID-19 testing, often performed using the polymerase chain ...

sequence at the

polyadenylation Polyadenylation is the addition of a poly(A) tail to an RNA transcript, typically a messenger RNA (mRNA). The poly(A) tail consists of multiple adenosine monophosphates; in other words, it is a stretch of RNA that has only adenine bases. In euk ...

site, also known as the poly(A) tail, and adds a second primer through the activity of

murine leukemia virus The murine leukemia viruses (MLVs or MuLVs) are retroviruses named for their ability to cause cancer in murine (mouse) hosts. Some MLVs may infect other vertebrates. MLVs include both exogenous and endogenous viruses. Replicating MLVs have a pos ...

reverse transcriptase A reverse transcriptase (RT) is an enzyme used to generate complementary DNA (cDNA) from an RNA template, a process termed reverse transcription. Reverse transcriptases are used by viruses such as HIV and hepatitis B to replicate their genomes, ...

. This permits reading full

cDNA In genetics, complementary DNA (cDNA) is DNA synthesized from a single-stranded RNA (e.g., messenger RNA (mRNA) or microRNA (miRNA)) template in a reaction catalyzed by the enzyme reverse transcriptase. cDNA is often used to express a speci ...

sequences and can deliver high yield from single sources, even single cells that contain 10 to 30

picogram To help compare different orders of magnitude, the following lists describe various mass levels between 10−59 kg and 1052 kg. The least massive thing listed here is a graviton, and the most massive thing is the observable universe. ...

s of mRNA, with relatively low levels (3-5%) of contaminating rRNA sequence. This technique is often employed in whole transcriptome shotgun sequencing. It is marketed by Clontech as Switching Mechanism At the 5' end of RNA Template (SMART) as well as by Diagenode as Capture and Amplification by Tailing and Switching (CATS).

Drop-Seq

By using syringe pumps to transmit a steady rate of isolated cells and uniquely oligonucleotide-barcoded beads, it is possible to isolate individual cells and beads together in droplets of lysis buffer, where the

site binds to a primer containing a unique, identifying sequence. This primer also contains a common sequence upstream of the identifier, so that after it is extended by reverse transcription, subsequent rounds of PCR will incorporate the tag, which permits each isolated cDNA that is sequenced to be tracked back to a specific originating bead. This permits the relative levels of transcripts in many individual cells to be analyzed simultaneously, creating a rational basis for the classification of these cells into particular cell types, or permitting the logical inference of in situ hybridization data from embryos without actually performing the experiment.

References

Reverse transcriptase inhibitors Polymerase chain reaction {{molecular-biology-stub