To delete the SNP rs55958994-containing enhancer-like region, 22Rv1 cells were transfected with plasmids containing Cas9 and guide RNAs targeting the enhancer-like region. Colonies were derived from single cells and tested for the loss of the target region. Cell clones were genotyped, and three clones with homozygous deletion of the rs55958994-containing region and three control clones were used for RNA-seq analysis. The single-guide RNA (sgRNA) information is listed in table S1, which was designed by using web-based tools at To generate site-directed mutation of SNP rs55958994, we transfected 22Rv1 PCa cells with plasmids containing Cas9, guide RNAs targeting the enhancer-like region, and DNA fragments containing (T) allele as repair templates. Double-nicking strategy was taken to reduce undesirable off-target mutagenesis. The sgRNA information is listed in table S4, which was designed by using web-based tools at The region containing mutation was amplified using specific primers (table S4), and SNP genotyping was performed by Sanger sequencing–based method.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.