shRNA/siRNA which loop sequence? - (Aug/07/2006 )
Again, many different statements to be found without a good explanation why this or the other hairpin loop sequence should be used. The promega shRNA (http://www.promega.com/siRNADesigner/program/default.asp), for example, offers 4 different presets for pGeneClip U1 vector, but a comparison is nowhere to be found.
suggested loop sequences for U1 hairpin from Promega shRNA designer website:
1) TTCAAGAGA (Brummelkamp, T.R.et al. , 2002)
2) AAGTTCTCT (Promega)
3) TTTGTGTAG (Scherr, M. et al., 2003)
4) CTTCCTGTCA (Schwarz D. S. et al., 2003)
Did you ever compare or read a comparison of loop sequences? Which vectors are you using with which loops?
All the best, j
We used brummelkamp's loop sequence and worked perfectly fine.
Nonsensey pointed out an interesting paper: Han2006. They analyse the cleavage of pri-miRNA depending on loop sequence & even test constructs where the loop has been cut open:
"the terminal loop is rather irrelevant to pri-miRNA processing. We note, however, that both processing and DGCR8 binding were slightly impaired in a mutant with a small loop"