Figure 2: Analysis of piggyBac mediated hSIRPA-BAC-TIR transposition. | Scientific Reports

Figure 2: Analysis of piggyBac mediated hSIRPA-BAC-TIR transposition.

From: Comparative Analysis of piggyBac, CRISPR/Cas9 and TALEN Mediated BAC Transgenesis in the Zygote for the Generation of Humanized SIRPA Rats

Figure 2

(A) Genotyping result showing PCR positive pups for a 201 bp regions in the human SIRPA gene. The diagram shows pup numbers 1.6, 4.2, 5.4, 5.5, 5.6, 5.7, 8.1, 9.2 and 9.4 to be positive for PCR. In the upper panel, BAC refers to RP11-887J4 used as a positive control, gDNA refers to human genomic DNA as a positive control, and Negative as a negative control. In the lower panel, 5.7 (Control) refers to genomic DNA from pup number 5.7 used as a positive control. (B and C) Show PCR result of rat zygotes injected with Cas9 mRNA transcript, instead of piggyBac transposase, as a negative control. BAC refers to RP11-887J4 used as a positive control. To determine transposition, splinkerette PCR was performed in order to sequence the region adjacent to the TIR ends. (D) Shows the sequencing result, indicating that six pups (1.6, 5.4, 5.7, 8.1, 9.2 and 9.4) carry transpositioned hSIRPA-BACs. The first column shows the pup identities, the second and third columns show the location of the transposition sites, the fourth column lists the TTAA site of transposition, and sequences ten base up and downstream. The last column indicates whether the transposition occurred in genic or intergenic region.

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