显微介导的远缘基因渐渗技术在鲤育种中的应用

Introgressive hybridization of exogenous gene by micro-injection incarp (Cyprinus carpio L.) breeding

以中国明对虾(Fenneropenaeus chinensis)为外源DNA供体,运用显微介导的远缘杂交技术,将中国明对虾总DNA直接导入鲤(Cyprinus carpioL.)受精卵内,创建鲤外源DNA导入系,并利用AFLP分子标记技术和肌肉营养成分分析,对显微介导中国明对虾基因鲤进行外源DNA检测与蛋白质和氨基酸含量进行测定,结果表明:(1)在30对AFLP引物中,有8对所扩增带,其显微介导中国明对虾基因鲤亲本和子代中都有和中国明对虾基因相同而对照鲤没有的带。说明受体鱼中,肯定有一段序列与目的基因序列一致,研究证明了亲缘关系甚远的异源DNA片段的超远缘杂交的可行性。(2)导入中国明对虾总DNA片段的显微介导中国明对虾基因鲤的蛋白质含量分别为18.37%、18.47%、17.99%,均高于对照组(17.06%);而氨基酸的总量分别为17.16%、17.56%、16.92%,也高于对照组(16.13%),其中天门冬氨酸、谷氨酸、甘氨酸、丙氨酸等4种鲜味氨基酸含量明显超过对照组,经数理统计分析,均有显著差异(P<0.05)。说明直接导入中国明对虾总DNA对显微介导中国明对虾基因鲤的营养成分和氨基酸含量均产生影响,这可对鲤的品质改进,丰富鲤的遗传基础,为显微介导的超远缘杂交技术的应用提供思路。

Introgressive hybridization is a movement of a gene from one species into the gene pool of another byrepeated backcrossing an interspecific hybrid with one of its parent species. It is an important source of geneticvariation in natural population. But the natural introgressive does not have the human direct interference while theexotic introgression is induced intentionally and very slowly. Transgenic technique is an extreme example ofintrogression verified as a successful hybridization from a transgenic plant to a wild relative. Carp is an importantspecies of fish, but has low muscle quality. The study that focused on the feasibility improves the quality ofmuscle of carp by introgressive hybridization technology. In this study, transgenic carp progenies were constructedby transferring all-fish gene of shrimp （Fenneropenaeus chinensis） to the zygote of common carp （Cyprinuscarpio L.） using mature micro-injection technology. AFLP technology was used to identify the transgenic zygote.The results of genotype of 30 pairs of AFLP markers and analysis of nutrient content of muscle showed that： （1）The gene of shrimp were detected in genome of transgenic carp by 30 pairs of primers of AFLP of shrimp whichconfirmed that the genomic fragment of shrimp exists in transgenic carp and has no found in contrast ones, thenindicates that it should be feasible to transmit genetic material between two distant related species by transgenicmethod at molecular level. （2） The statistic analysis showed that the contents of crude protein of transgenic carpswere 18.37%, 18.47% and 17.99%, which were higher than those in non-transgenic ones（17.06%）. Total aminoacids contents were 17.16%, 17.56% and 16.92% respectively, also higher than that of control （16.13%）, of which4 kinds of amino acids contents in transgenic carp related to taste were higher than those in common carpsignificantly（P0.05）, such as aspartic, glutamic, glycin and alanine. So, the effects of shrimp gene directlytransmitted to the carp genome gives rise to increase nutrient content and amino acids. This method would beuseful in improving carp muscle quality and heredity base, and was a very powerful technology for exogenousDNA transferred between distant related species. In the future, more and more good trait of fish or other specieswould be improved by this technology, although this technique needs developing further. Transgenic fish will beused in farm after it is tested no harm to ecosystem and human beings.