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  • Generation of a homozygous CIITA knockout iPS cell line using the CRISPR-Cas9 system.

Generation of a homozygous CIITA knockout iPS cell line using the CRISPR-Cas9 system.

Stem cell research (2021-10-24)
Elena Romano, Piera Trionfini, Roberta Giampietro, Ariela Benigni, Susanna Tomasoni
ABSTRACT

Human induced pluripotent stem cells (iPSCs) have great promise in regenerative medicine. However, several limitations, including immune-incompatibility, have raised concerns regarding their clinical application. Recent studies have shown that human iPSCs and their derivatives lose their immunogenicity when major histocompatibility complex (MHC) class I and II genes are inactivated and CD47 is over-expressed. In this study, we used CRISPR-Cas9 technology to generate an isogenic iPSC line with a homozygous frameshift mutation in the MHC II transactivator (CIITA) gene. The CIITA-/- iPSCs exhibit typical morphology of pluripotent cells, normal karyotype, expression of pluripotency markers and differentiation capacity in the three germ layers.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-TRA-1-81 Antibody, clone TRA-1-81, clone TRA-1-81, Chemicon®, from mouse
Sigma-Aldrich
Anti-Actin, α-Smooth Muscle - Cy3 antibody, Mouse monoclonal, clone 1A4, purified from hybridoma cell culture
Sigma-Aldrich
Anti-Tubulin β-III Antibody, clone TU-20, Alexa Fluor488 Conjugated, clone TU-20, from mouse, ALEXA FLUOR 488
Sigma-Aldrich
DAPI, for nucleic acid staining
Sigma-Aldrich
Anti-TRA-1-60 Antibody, clone TRA-1-60, clone TRA-1-60, Chemicon®, from mouse