| Trachinotus blochii is the coastal areas of South China the important mariculture fish.Because of the delay of the breeding work,cultured Trachinotus blochii face decreases production performance problems.Breeding of fine varieties Trachinotus blochii,is one of the important means to solve the problem.The molecular marker assisted breeding and traditional breeding combined strategy can greatly improve the breeding efficiency in Trachinotus blochii.Insulin like growth factor gene IGF has the function of regulating cell metabolism,promoting cell growth and inhibiting cell apoptosis.It is a major growth factor,which plays an important role in the growth and reproduction of fish.It is proved to be an important molecular marker gene for growth traits.So this thesis to Trachinotus blochii as the research object,to carry out correlation analysis of Trachinotus blochii groups in IGF-I gene gene and growth traits.1.Applying Race technology to colon the IGF-I gene cDNA sequence of Trachinotus blochii and get a new cDNA sequence with a length of 1680bp,the 5’UTR is 123bp,open reading frame(ORF)is 558bp and encoding 185 amino acids.The results show that Trachinotusblochii IGF-I protein have a signal peptide,which carries 43 amino acids and its protein structure contains three reverse alpha helix.Through homologous analysis and phylogenetic tree analysis,it is revealed that there is high homology between Trachinotus blochii IGF-I gene and percid IGF-I gene.2.By taking advantage of IGF-I gene cDNA sequence of Trachinotus blochii and amplifying the exon and intron sequences of segments of IGF-ITrachinotus blochii,5 exons and 3 introns are detected.The size of exons and intros respectively are:147bp for exon 1,1366bp intron 1,141bp for exon 2,176bp for exon 3,762bp for intron 3,29bp for exon 4,1366bp for intron 4,52bp for exon 5.Growth traits such as length,weight,height of three Trachinotus blochii group are measured and then the IGF-I gene is taken into SNP analysis.Three SNP loci are detected(A161G,T172C,T174C):the A-to-G mutation of the 161 base in intron 1(A161G),two T-to-C mutations of the 174 and 199 base in intron 3(T172C,T174C).The results show that the SNP loci of IGF-I gene associated with growth traits are not found in 3 groups.The eggs of Trachinotus blochii IGF-I gene in Hainan group may be quite conservative.3.Unable to find Trachinotus blochii IGF-I gene and growth related gene type,this study cloned gf-i gene sequences upstream regulatory elements were analyzed.By chromosome walking shift method for cloning Trachinotus blochii IGF-Ⅰ gene promoter sequence for 1195bp,and respectively and largemouth bass,zebrafish,sheep IGF-Ⅰ start subsequence blast comparison,both similarity respectively 90%,86%and 88%.Using software of Trachinotus blochii IGF-Ⅰ gene start subsequence potential transcription factor binding sites were analyzed and sequence prediction analysis shows that,the promoter has nine potential transcription factor binding sites,(SP1,HNF-3,STST5b,GR,ARP-1,GR,ER,C/EBP),in order to further reveal the regulatory mechanism of Trachinotus blochii IGF-Ⅰ gene laid the foundation. |
