精子介导鱼类基因转移和聚合酶链反应检测技术

SPERM MEDIATED GENE TRANSFER AND METHOD OF DETECTATION OF INTEGRATED GENE BY PCR

金鱼精子与美洲大绵■的抗冻蛋白基因一起保温３０分钟后，再与卵子受精，共获得１４５尾成鱼和若干胚胎。从胚胎和成鱼中提取 ＤＮＡ经聚合酶链反应（ＰＣＲ法）扩增和 Ｓｏｕｔｈｅｒｎ ｂｌｏｔ分子杂交表明，外源的抗冻蛋白基因进入了部分受体鱼的染色体组内。测定的４５尾一年龄实验鱼中，有１２尾显示出明确的杂交带，阳性率为２６％。

Sperm Of goldfish were incubated with antifreeze protein gene at 4℃ for 30min. Then, they were mixed with eggs for fertilization. Total 145 experimental fish(one-year old)and a lot of embryos were obtained. DNA was extracted from red cells of the experimental goldfish. Forty-five samples were analyzed by PCR and Southern blot using 2 OKb-fragment of AFP gene as the probe. The results showed that DNA from both embryos and adult goldfish showed clear bind of hybridization. The results indicated that foreign AFP gene were indeed transfered into the goldfish eggs. Twelve of the 45 samples were found positive,and the positive rate was 26%.