In this study, we describe the application of nucleotide isomers that form non-standard base pairs in the template switching oligo to prevent background cDNA synthesis. When such bases are added to the 5' end of the template switching (TS) oligo, they inhibit MMLV-RT from extending the cDNA...
Group II intron RTs have an end–to–end template-switching activity similar to that of MRP and R2 element RTs (6, 39), and this activity has been utilized for adapter addition in RNA-Seq using a thermostable group II intron RT (TGIRT-seq) (6, 8, 9). Unlike the R2 element RT (...
This method is relatively rapid, but poly(dT) tailing and variable non-templated nucleotide addition by the retroviral RT to the 3′ end of the DNA product prior to template switching make it difficult to precisely identify the DNA template ends and can introduce strong bias for regions with l...
The ligation reactions (15 μl) were overnight at 4 °C in T4 RNA ligation buffer, 1 mm ATP, 20 units of ligase, 3.4 pmol of 177-nt RNA, and 5 pmol of CG or 5.9 pmol of AT RNA. Products of the ligations were RT-PCR-amplified, SpeI-digested, and analyzed as described above....
Leave ample space or buffer time in your calendar to stop yourself from getting derailed. Alternatively, try to always overestimate the time you think you’ll need to complete a task — at least until you’ve done it enough times to be confident about how much time it really takes. ...
A nontemplated-addition reverse transcription reaction (no TSO present; 30-μl total volume) contained 1 μm RNA template (30 nmol), 1× Template Switching RT Buffer (catalog number B0466), 2 mm dNTP solution mixture (catalog number N0447), 0.5 μm 5′-FAM V5 primer, and 400 units of...
(PGE) with AuNPs, free radical polymerization was conducted in the presence of N-acryloyl pyrrolidine-2,5-dione (NAPD, functional monomer), EGDMA (cross-linker), AIBN (initiator), D- and L-aspartic acid (templates) and MWCNTs, followed by template removal with NaOH and phosphate buffer. Al...