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Asian-Australas J Anim Sci > Accepted
doi: https://doi.org/10.5713/ajas.16.0697    [Accepted]
Human lactoferrin efficiently targeted into caprine beta-lactoglobulin locus with TALENs
Yu-Guo Yuan1,2, Shao-Zheng Song1, Meng-Ming Zhu1, Zheng-Yi He1, Rui Lu1, Ting Zhang1, Fei Mi1, Jin-Yu Wang1,*, Yong Cheng1,2,*
1Yangzhou university, Yangzhou, China
2Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou, 225001, China
* Corresponding Author: Jin-Yu Wang ,Tel: +86-514-87979348, Fax: +86-514-87972218, Email: cheng1391@hotmail.com / Yong Cheng ,Tel: +86-514-87979348, Fax: +86-514-87972218, Email: cheng1391@hotmail.com
Received: September 19, 2016;  Revised: November 11, 2016.  Accepted: December 13, 2016.  Published online December 17, 2016.

ABSTRACT
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.
Keywords: Beta-lactoglobulin; Gene targeted; TALENs; Human lacotoferrin; Goat; Nuclear transfer
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