Production of Transgenic Chimeric Chicken from Cryopreserved Primordial Germ Cells and its Validation by Developing shRNA Transgenic Chicken Chimera

Primordial germ cells (PGCs) are precursors of gametes in birds. For ex-situ conservation and production of transgenic birds, there is a limitation for preservation of oocytes in birds as compared to other mammalian species. To overcome those limitations, PGCs have been used as candidate cells, which have been cryopreserved and manipulated and used as to produce transgenic birds. In this study, cryo-preserved PGCs were used to produce transgenic birds. The protocol for production of transgenic birds with cryo-preserved PGCs was developed and the success rate for production of transgenics 16.7% in the protocols established in the study. The same gene transfer protocol through PGCs was validated by transferring shRNA molecule of SREBP-1 gene to the host genome to produce transgenic chimeric birds and the success rate for production of transgenic chimeric chicken was 40%. Finally, it is concluded that a standard protocol for ex-situ conservation of birds through PGCs and production of transgenic birds from cryo-preserved PGCs and knock down birds from PGCs were developed. It may be suggested that these protocols for resurrecting live birds from cryo-preserved PGCs may be applied as model for protecting the endangered birds from their extinction.