Received: September 19, 2016; Revised: November 11, 2016. Accepted: December 13, 2016. Published online December 17, 2016.
Objective: To create genetically modified goat as a biopharming source of recombinant human lacotoferrin (hLF) with TALEN.
Method: TALENs and targeting vector were transferred into cultured fibroblasts to insert hLF cDNA in the goat beta-lactoglobulin (BLG) locus with homology-directed repair (HDR). The gene targeted efficiency was checked using sequencing and TE7I assay. The bi-allelic gene targeted colonies were isolated and conformed with PCR, and used as donor cells for somatic cell nuclear transfer (SCNT).
Results: The targeted efficiency for BLG gene was approximately 10%. 12 bi-allelic gene targeted colonies were isolated, five of which were used in first round SCNT and 4 recipients (23%) were confirmed pregnancy at 30 d. In second round SCNT, 7 (53%), 4 (31%), and 3 (23%) recipients were confirmed to be pregnant by ultrasound on 30 d 60 d, and 90 d.
Conclusion: This finding signifies the combined use of TALENs and SCNT for generation of bi-allelic knock-in fibroblasts and cloned fetus. Therefore, it might lay the foundation for transgenic hLF goat generation and mammary gland as a bioreactor for large-scale production of recombinant hLF.
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