New Delhi, Sep 08, 2010: After producing two cloned buffalo calves, Haryana-based National Dairy Research Institute (NDRI), India, through the new and advanced ‘Hand-guided Cloning Technique’, has cloned another buffalo calf, named Garima-II on August 22, 2010. This cloned buffalo calf is different from the earlier cloned calf because, in this case, the used donor cell was embryonic stem cell.
The world's first buffalo calf through the “Hand guided Cloning Technique” was born on February 6, 2009 at NDRI, Karnal and subsequently, the second cloned calf “Garima”, weighing 43kgs, was born on June 6, 2009 . In earlier cloning, the donor cell was from somatic cells. The donor embryonic stem cell was isolated from the eight-day old blastocyst. These cells were cultured up to 29-passages (117 days) till it expressed pluripotent marker and then confirmed to be stem cell.
Dr AK Srivastava, Director of the institute emphasized that this technology could go a long way in helping faster multiplication of superior milch buffaloes in India. “Although the world’s largest population of buffaloes is in India and they are contributing about 55 percent of total milk production in country, but the percentage of elite animals is very low and there is an urgent need to enhance the population of these elite buffaloes,” he said. He further emphasized that there is an acute shortage of good bulls and the technology of cloning will decrease this gap between supply and demand of breeding the bulls in the shortest possible time.
The team involved in the production of this cloned calf using embryonic stem-cell as donor cell are Dr MS Chauhan, Dr SK Singla, Dr RS Manik, Dr P Palta, Dr Shiv Parsad, and Dr Aman George of NDRI, Karnal, Haryana. The scientists are of the opinion that the embryonic stem cells have better cloning ability as compared to somatic cells and that the epigenetic reprogramming of these cells is much more efficient than the somatic cells, which are already differentiated and lineage committed.The scientists believe that the cryopreservation of embryos will need to be made as part of technique, so that the embryos could be transported and used at several places.
The hand-guided cloning technique developed at NDRI, is an advanced modification of the “Conventional Cloning Technique”. In this technique, immature oocytes were isolated from ovaries and were matured in vitro. These were then denuded and treated with an enzyme to digest the outer layer of oocytes called ‘zona pellucida’. The oocytes were then treated with chemicals to push their genetic material to one side of the oocyte. This protruded side was then cut off with the help of “hand held fine blade” for removing the original genetic material of the oocyte. The enucleated oocyte was then electrofused with single cell taken from colony of embryonic stem cells. The resulting embryos were cultured and grown in the laboratory for seven days to develop them to the stage of blastocyst. The blastocysts were transferred to recipient buffaloes.
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