Isolation Of Ghrs, Igf-Ⅰ Genes And Snps Associated With Growth Traits Gift Strain Orechromis Niloticus

Animal growth was mainly regulated by the growth axis of growth hormone (GH)/insulin-like growth factor (IGF). Growth hormone receptors (GHR) is one of important factors for the growth axis. Growth hormone (GH) interacts wiht GHR on tissues to evoke post-receptor signaling events. So the mutations on growth hormone receptor genes play a vital important role on animal growth. Two cDNA sequences and DNA sequences of exon2-9of GHR1, exon2-8of GHR2were isolated from GIFT strain (Genetically Improved Farmed Tilapia strain) using RT-PCR and PCR in this study. And we identified six and sixteen SNPs on GHR1and GHR2respectively, only two of which were located at exons. PCR-RFLP was performed to detect genotypes in eight SNPs (GHR1intron3_A612G, intron3_A989T, intron7_A599C; GHR2intron3_C330T, intron3_A645G, intron3_G687T, intron3_A967G, intron7_C107T) of120individuals from five families of GIFT strain. Correlation analysis between genotypes and weight gain showed that GHR1intron3_A612G and A989T associated with male fish weight gain (P<0.01and P<0.05) significantly. The weight gain of heterozygote was the most. Similar situation was found in the female population, but the difference was not significant (P>0.05). Intron7_A599C was significantly associated with both male (P<0.01) and female (P<0.05) fish weight gain. The CC type fish weight gain was significantly higher than AA type’s. On the GHR2, only intron3_G687T was found to correlate with female weight gain significantly (P<0.05). The GT type female fish gains was significantly higter than GG type’s. In order to validate the performance of screened markers in other families, the correlation between genotypes of A612G, A989T, A599C and G687T and weight gain were detected in356individuals from60families of GIFT strain (6individuals in average of each family). The diverse genetic background and smaller number of samples result in the decreased significance, but there were similar growth situation among different genotypes’fish. And the four molecular markers identified in this study can be used as a reference in GIFT strain breeding. Real-time quantitative PCR results indicated that GHR1expression in male fish livers was1.36times higher than that in female’s, and except in kidney, the male fish GHR2expression in other tissues was lower than the female, which can explain the different effects of GHR1and GHR2SNP loci on the weight gain of male and female GIFT. In addition, the different influences of GHR1and GHR2on the growth result in only one SNP loci on GHR2could affect the weight gain.Insulin-like Growth Factor1(IGF-I) is also one of important genes controlling growth, development in animals. Compared the exonl-partial intron2-exon3-partial intron4sequences of IGF-I from6individuals of Genetically Improved Farmed Tilapia strain (GIFT strain), we identified3SNPs on jfIGF-I in this study: intron1_A1307G, intronl_G1316T and intron3_C6T. PCR-RFLP was performed in intron1_A1307G and intron3_C6T genotypes detection of121individuals from five families of GIFT strain. Results of the correlation between genotypes and weight gain, body form showed that intron1_A1307G was significantly associated with female fish weight gain and ratio of body thickness/length (P<0.05). The weight gain and thickness/length value of AA type female fish was significantly higher than that of GG type. However, the correlation with the male fish was not significant (P>0.05). Intron3_C6T was significantly associated with both male (P<0.05) and female (P<0.01) fish weight gain. The weight gain of TT and CT type male fish was significantly higher than CC type; the weight gain of female with the CT genotypes was significantly higher than TT. The intron3_C6T was also significantly associated with female fish height/length value (P<0.05). The height/length value of the TT and CT genotypes female fish was significantly higher than CC. In order to validate the performance of screened markers in other families, we detected the correlation between intronl A1307G, intron3_C6T genotypes and weight gain among417individuals from60families of GIFT strain (7individuals in average of each family.) And similar results were observed. The diverse genetic background and smaller number of samples resulted in the decreased significance. These results showed that the relationship between genetic background and the number of samples should be considered during molecular makers screening. And the two molecular markers intronl_A1307G, intron3_C6T, identified in this study can be used as a reference in GIFT strain breeding.