Fig. 5: Establishment of TYRP1 modified PGC lines using the CRSPR/Cas9 system. | Communications Biology

Fig. 5: Establishment of TYRP1 modified PGC lines using the CRSPR/Cas9 system.

From: PGC-based cryobanking, regeneration through germline chimera mating, and CRISPR/Cas9-mediated TYRP1 modification in indigenous Chinese chickens

Fig. 5

A Schematic representation of the TYRP1 gene structure on chromosome Z and the nucleotide sequence of the exon 2 target region, which contains four target sites (sgRNA#1–#4). The four sgRNA sequences and the protospacer adjacent motif (PAM) sequences are underlined in red and light green, respectively. The two black underlined primers were used to amplify the target region (371 bp). B PCR products for the exon 2 target region using each bulk DNA sample from TYRP1 modified PGC lines using three sets of sgRNAs (#1/#2, #1/#3, and #1/#4) (left), along with cleavage efficiency assessed by the T7E1 assay (right). C Sanger sequencing analysis of TA clones of the target region for the TYRP1 modified PGC line (#CA10_TYRP1-KO) using one sgRNA set (#1/#2). Sequence chromatograms and partial sequences show two representative CRISPR/Cas9-induced mutations: 73 bp deletion (frameshift mutation) (left) and 74 bp deletion (frameshift mutation) (right) in the target region. The sequences of the sgRNAs are underlined in red and the PAM sites are underlined in light green. Red arrows indicate induced mutations.

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