Warning: imagejpeg(C:\Inetpub\vhosts\kidney.de\httpdocs\phplern\26933249
.jpg): Failed to open stream: No such file or directory in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 117 Cold+Spring+Harb+Protoc
2016 ; 2016
(3
): pdb.prot090803
Nephropedia Template TP
gab.com Text
Twit Text FOAVip
Twit Text #
English Wikipedia
Single Guide RNA Library Design and Construction
#MMPMID26933249
Wang T
; Lander ES
; Sabatini DM
Cold Spring Harb Protoc
2016[Mar]; 2016
(3
): pdb.prot090803
PMID26933249
show ga
This protocol describes how to generate a single guide RNA (sgRNA) library for
use in genetic screens. There are many online tools available for predicting
sgRNA sequences with high target specificity and/or cleavage activity. Here, we
refer the user to genome-wide sgRNA sequence predictions that we have developed
for both the human and mouse and that are available from the Broad Institute
website. Once a set of target genes and corresponding sgRNA sequences has been
identified, customized oligonucleotide pools can be rapidly synthesized by a
number of commercial vendors. Thereafter, as described here, the oligonucleotides
can be efficiently cloned into an appropriate lentiviral expression vector
backbone. The resulting plasmid pool can then be packaged into lentiviral
particles and used to generate knockouts in any cell line of choice.
free
free
free
