From 9214d2093c09f79904e94c0911a15a937d6006da Mon Sep 17 00:00:00 2001 From: Charles Plessy Date: Thu, 19 Apr 2018 13:45:24 +0900 Subject: [PATCH] More details on Mg / Mn addition. --- tags/template_switching.mdwn | 16 ++++++++++++---- 1 file changed, 12 insertions(+), 4 deletions(-) diff --git a/tags/template_switching.mdwn b/tags/template_switching.mdwn index 7af520cb..f24e5178 100644 --- a/tags/template_switching.mdwn +++ b/tags/template_switching.mdwn @@ -12,10 +12,6 @@ can extend a linker with the sequence of a small RNA via a template switching reaction. (That is: a sRNA can play the same role as a TS oligonucleotide.) - - [[Lee et al (2017)|biblio/28327113]] increased the efficiency of template - switching non-capped molecules by increasing dNTPs to 2 mM and - Mg2+ to 9 mM. - ### Effect of chemical composition of the TS oligonucleotide Originally, the TSOs were all-RNA. Since this is expensive to synthesise, @@ -33,4 +29,16 @@ as a replacement for RNA. - [[Arguel et al (2017)|biblio/27940562]] reported similar performance for RRR and RRL, using a 5′-focused method similar to nanoCAGE or STRT. +### Effect of magnesium, manganese and dNTP concentrations + + - [[Schmidt and Mueller, 1999|biblio/10518626]] showed that increasing + magnesium concentration (to 6 mM) or adding manganese at the end of the + reaction (1 or 2 mM) increased the frequency of dC addition (moderately + for Mg2+ and strongly for Mn2+). Enzyme: SSII; dNTP + concentration: 1 mM each. + + - [[Lee et al (2017)|biblio/28327113]] increased the efficiency of template + switching non-capped molecules by increasing dNTPs to 2 mM and + Mg2+ to 9 mM. + [[!inline pages="tagged(template_switching)" limit=0]] -- 2.47.3