金鱼Tgf2转座子插入位点多态性鉴定及其对关联基因结构和表达的影响

Goldfish, originated from the crucian carp in lower reaches of the Yangtze River, possesses hundreds of varieties. According to the phenotypes, goldfish are divided into three linages, i.e., grass, wen and egg. Though goldfish are so different from common crucian carp in morphology, it belongs to the same species. Our previous studies have shown that the goldfish genome contains an active DNA transposon, Tgf2, which is not detected in the crucian carp. Given transposon serves as a main source of genetic variation, it is speculated that the Tgf2 transposon might played important roles in genetic differentiation of goldfish. In the present study, Tgf2 transposon insertion polymorphism (Tgf2-TIP) identification will be conducted using high throughput sequencing technology, followed by comparative analysis among the three linages for determination of the shared or specific Tgf2-TIPs. Then, the TIP associated genes will be annotated according to the zebrafish and common carp genome followed by exploring the genetic effect of the insertion of Tgf2 on gene structure and expression by gene cloning, qPCR and in situ hybridization. Finally, RNAi and gene knock out will be conducted to elucidate the function of the genes in embryonic development and post-embryonic morphogenesis in zebrafish and crucian carp. The study will help in revealing the genetic basis of rapid adaptive evolution of goldfish under artificial selection.

金鱼是鲫鱼的一个分支，在人工选择下发生快速进化，产生了丰富的表型变异。研究表明，金鱼为单系起源，但其快速进化的遗传基础仍不明确。申请人的前期研究表明，金鱼基因组中存在高活性的Tgf2转座子，而鲫鱼基因组中不存在，推测Tgf2转座子可能在金鱼快速进化中发挥了重要的驱动作用。本申请项目利用转座子插入富集文库测序技术鉴定金鱼基因组中Tgf2的插入位点多态性，进而在三个品系间进行比较分析，筛选品系间共有和品系（品种）内特有的Tgf2插入位点。以斑马鱼和鲤鱼基因组为参考，对插入位点的上下游基因进行功能注释，利用基因克隆、qPCR和原位杂交技术研究Tgf2转座子对关联基因结构和表达的影响；以斑马鱼和鲫鱼为模型，利用RNAi和基因敲除技术模拟Tgf2转座子插入对关联基因结构和表达的影响，研究关联基因在胚胎发育和形态建成中的功能。研究结果将为解析金鱼快速进化的遗传机制提供新的研究思路和内涵。