Analysis Of DNA Molecular Markers With The Candidate Genes Of Economic Traits In Paralichthys Olivaceus And Larimichthys Crocea Hatchery Stocks

Polymorphisms in economic trait candidate genes and theirs association with economic traits in different olive flounder Paralichthys olivaceus and large yellow croaker Larimichthys crocea hatchery stocks were analyzed by molecular marker technology.Based on the PCR-SSCP analysis of GH gene in Jiaonan flounder hatchery stock, the exon4 was shown to be polymorphism and had one substitution mutation at site 1763 (c→t) in AB genotype. The evaluation of the association between SSCP patterns with weight trait and head lengthen suggested a positive effect of pattern AB of exon4 (P<0.05) by SPSS. Allele B was an effective factor for flounder weight and head lengthen. MANOVA test revealed that flounder in different genotype had different body shape. Chi-square test showed that this polymorphism locus in the stock was in Hardy-Weinberg equilibrium (P<0.05). In the GH exon1 region, a microsatellite locus was detected. Five genotypes and three alleles were tested. One-way ANOVA result showed that fish with genotype AC were heavier than fish with other genotypes and C was an effective allele in flounder weight.The SSCP study in Larimichthys crocea from two different locations showed GH gene in the Zhejiang stock exhibited one SNP at position 196 (g→a). Heterozygous genotype had negative effect on body height while positive effect on standard length / body height (P≤0.05). This result indicated that this locus could be used as a candidate marker for large yellow croaker body shape. Polymorphism of GH gene in the Fujian stock was also discovered due to SNP at position 692 (t→c). Genotype CD had significantly positive effect on both weight and total length (P≤0.01). It also hinted us that this mutant maybe a weight trait and head length candidate marker in large yellow croaker.Genetic diversity of two cultured stocks of Paralichthys olivaceus from Jiaonan County and Rizhao County were analyzed by using one microsatellite locus in GHR gene, and the polymorphisms and their associations with growth traits were also investigated. Their allele numbers were 12 and 9, respectively, the effective alleles numbers were 6.26 and 5.04. The chi-square test presented that this microsatellite locus in both stocks were deviated from the Hardy-Weinberg equilibrium (P<0.01). ANOVA analysis suggests the individuals with genotype IM was highest in all morphological values of the Jiaonan stock but had only significantly difference in weight with all other genotypes (P<0.01). While in the Rizhao stock, values of weight, total length, body height, tail height, tail length and distance between eyes of the individuals with genotype BC were higher than those with other genotypes, and values of body length and head length of the individuals with genotype CJ were higher than those with other genotypes. Genotype IM in Jiaonan Paralichthys olivaceus could be used as a candidate marker in weight.In order to select molecular markers with anti-disease trait, 185 healthy olive flounder were challenged with Edwardsiella tarda LSE40. Death group and survival group of olive flounder were obtained. TLR2, TRL3 and TLR9 genes were used as candidate genes and 7 pairs of primer were designed to amplify parts of these genes sequences. RFLP was applied in analyzing the polymorphism locus. After primary screening, the PCR product amplified by the primer TRL3C was found to exist a SNP caused by a mutant in site 3806 of TLR3 gene. This mutant resulted in absence of EcoRI restriction site in some individuals. Three different patterns, named as AA, AB and BB, were presented in the EcoRI restriction enzyme map. Chi-square test proved that this variance was associated with the flounder anti-disease performance. Logistic analysis was carried through flounder growth traits and the result indicated that head length, body length and body height had significant difference (P<0.05) while they had no relativity with weight. Head length and weight were selected into Logistic analysis and result revealed that genotype AA had notable impact on death (P<0.05). AA was the most dangerous factor while AB had no significant effect on death. Head length was the major protect factor (P<0.05) while weight had no significant effect on death (P>0.05). A was the most dangerous allele for death while B was the most favorable allele for survival verified by Chi-square test. The result indicated that this locus might be considered as a potential marker for olive flounder anti-disease.