CRISPR/Cas9 constructs
sgRNAs targeting promoters of genes of interest
(A3A , A3B , AID , A3C , A3D , A3H )
were designed using the open-access web tool
Chop-Chop65. PCR products containing the U6 promoter
and sgRNA specific for every promoter region were synthesized by
two-step mutagenic PCR using Q5 polymerase and purified using a Qiagen
gel extraction kit. The following plasmids were used: pcDNA-dCas9-p300
Core (AddGene plasmid #61357) and pcDNA-dCas9-p300 Core (D1399Y), gifts
from Dr. Charles Gersbach; pLX-sgRNA (AddGene plasmid #50662), a gift
from Dr. Eric Lander and Dr. David Sabatini; dCas9-p300 Core was cloned
into Lenti-Cas9-2A-Blast plasmid (AddGene plasmid #73310; a gift from
Dr. Jason Moffat) instead of Streptococcus pyogenes Cas9.
dSaCas9-p300 was generated from pJEP303-pAAV-CMV-dSaCas9-VP64-pA
(AddGene plasmid #113678; a gift from Dr. Jonathan Ploski) by cloning
dSaCas9 into pcDNA-dCas9-p300 Core plasmid. dSaCas9 sgRNAs were produced
from a pSaGuide (Addgene plasmid #64710; a gift from Dr. Kiran
Musunuru). All primers are listed
in Table S1.