Right here, the brand new UTF1 supporter/enhancement factors manage EGFP phrase, as the puromycin resistance cassette try constitutively expressed immediately after directed genomic integration (Figure 4A). Co-transfection of target vector pattP4X-PGKssPuro-UTF1-EGFP and you will Int-C3NLS phrase vector followed closely by possibilities with puromycin contributed to 120 territories. Original PCR examination using ORF1 submit primer followed closely by PCR tool sequencing known 17 clones that features the recombination junction attL, proving successful attH4X focusing on (investigation not shown). Each other attL and you will attR junctions were verified in the 5/17 clones by the genomic PCR and sequencing, and clones A3, E3 and you may K3 was chose by the intergenic chromosomal location away from transgenes and the number of EGFP phrase. (Profile 4B; studies perhaps not found; Supplementary Dining table S5). (more…)
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