Genetic Analysis Of Key Function Genes And Their Associations With Economic Traits In Goats

The bioinformatics, DNA sequencing, DNA sequence analysis, PCR-SSCP, PCR-RFLP and AS-PCR techniques were applied to not only clone and predict the goat POU1F1 gene, but also scan and analyze the genetic variations of key function gene and their association with economic trais in 1384 goats, which comprised of ten breed, namely, Xinong Sannen dairy goat (Sa, n=74), Guanzhong dairy goat (GZ, n=62), Laoshan dairy goat (LS, n=80), Inner Mongolia white cashmere goat (IMWC, n=847), Northern Shaanxi white cashmere goat (NSWC, n=213), Banjiao goat (BJ, n=25), Matou goat (MT, n=22), Guizhou white goat (GW, n=31), Guizhou black goat (GB, n=21) and Leizhou goat (LZ, n=34). The five key function gene included POU1F1 gene (exon 1～6, intron 1, 3～5, 5'UTR and 3'UTR), PRL gene (exon 1～5), IGFB3 gene (exon 1～3 and intron 1,2), LALBA gene (exon1, 3, 4),CSN1S1 gene. The economic traits comprised of milk yield, wool yield, wool thickness, wool length, litter sizes, birth weight, weight, and body sizes). The association between genetic variations and economic traits were analyzed by using general linear model (GLM) or adjusted linear model, as well as population genetic structure and distribution of genotype and allele were calculated by chi-square and quantitative genetic method. These will benefit for the protection, development and application of goat genetic resource, application of DNA marker related to economic traits on marker-assist-selection (MAS), and improvement and promotion of dairy goat, meat goat and wool goat. The important results were in the followings:1 The clone and bioinformatics prediction of goat POU1F1 geneThe goat POU1F1 gene, a 876 bp length coding region (CDS), coded 291 amino acid and was organized by exon 1(No.1～142nt, 142bp), exon 2(No.143～214nt, 72 bp), exon 3 (No.215～439nt, 225 bp), exon 4 (No.440～604nt, 165 bp), exon 5 (No.605～665 nt, 61 bp) and exon 6 (No.666～876 nt, 211 bp), as well as intron 4 (747 bp). The prediction demonstrated the theoretical pI with 8.36, the predominant second protein structure with helix, the transmembrane region with No.63～79 amino acid, POU domain with the No.124～198 amino acid and homology domain with No.215～271 amino acids. The goat POU1F1 gene and its amino acids showed the highest homology with sheep, respectively, next is bovine. 2 Genetic variations of goat POU1F1 geneThe eighteen genetic variations were found at exon 3～6, intron 1, 3,4 and 3'UTR loci in goat POU1F1 gene : IVS1+10G>T,EX3₁13G>T,EX3₁14G>T,IVS3+8C>T, IVS3+41T>G, IVS2+41-42InsT, IVS3+175T>A, EX4₅3delA, IVS4+318T>A, VS4+383G>A, IVS4+710T>G, EX5₁4G>A, EX5₃4G>A, EX5₅9G>A, EX6₅8T>C, EX6₁72T>C, 3'UTR+92G>A, 3'UTR+92T>C. Among them, two variations were in exon 3, as well as one for exon 4, three for exon 5, two for exon 6, one for intron 1, four for intron 3 and intron 3, respectively, two for 3'UTR). In these mutations, ten were in non-coding region and eight in coding region (including three missense mutations, three silent mutations, one deleletion and one nonsense mutation). Among them, EX3₁13G>T resulted in 109Glu>His in protein, as well as EX3₁14G>T in 110Glu >X (Stop codon), EX4₅3delA in 164Glu shift frame, EX5₁4G>A in 206Glu>Glu, EX5₃4G>A in 213Arg>Lys, EX5₅9G>A in 218Arg>Lys, EX6₅8T>C in 241Ser> Ser, EX6₁72T>C in 279Ser>ser. Besides, IVS4+383G>A and IVS4+710T>G resulted in the polymorphism of IVS4 with wild and mutation type.3 Spices, AluI and DdeI PCR-RFLP of goat POU1F1 geneEX3₁14G>T resulted in the absence of No.111-291 peptide and maybe produce"the imagined alpha-POU1F1 spice"; EX4₅3delA resulted in the shift frame from No.164 AA in POU1F1 protein and may produce"imaginedδ-POU1F1 spice"; EX6₁72T>C mutation produced AuI polymorphism: allele C with 216 bp, 124 bp and 110 bp fragments, allele T with 340bp and 110 bp fragmet;EX6₅8T>C produced DdeI polymorphism: D1D1 genotype with 200bp, 118 bp, 102 bp, 20 bp and 11 bp, and D1D2 genotype with 200bp, 118 bp, 113 bp, 102 bp, 20 bp and 11 bp.4 Associations of genetic variations with economic traits in goat4.1 Associations of genetic variationof Ex1 locus with economic traits in goatThe polymorphism of Ex1 locus (including exon 1 and intron 1) was resulted by IVS1+10G>T. In this locus, the genotype GG, GT and TT were detected in analyzed populations and the predominant GG genotype and G allele presented in most populations except Xinong Sannen dairy (Sa), Guanzhong (GZ), Laoshan (LS) and Inner Mongolia white cashmere (IMWC) goat. The PIC values varied in 0.086～0.3362. It was significantly found that the distribution of genotypes and alleles associated with characteristics of breeds and goat types (dairy type, meat type and wool type) (P<0.05, P<0.01). The correlations were observed between this locus and litter size (P<0.05): the first birth litter size of individuals with genotype GT were significant higher than that of genotype GG (P<0.05). It implied that Ex1 locus had significant effect on reproduction performance.4.2 Associations of genetic variations of Ex3 locus with economic traits in goatThe polymorphism of Ex3 locus (including exon 3 and intron 3) was resulted by EX3₁13G>T, EX3₁14G>T, IVS3+8C>T, IVS3+41T>G, IVS3+41-42insT and IVS3+175T>A. The genotype GG, GA and AA were detected in analyzed populations and the predominant GG genotype and G allele presented in analyzed populations. The PIC values varied in 0.000～0.1555. Distribution of genotypes and alleles were founded to be significantly associated with characteristics of breeds (P<0.05, P<0.01). The correlations were observed between this locus and milk performance and wool performance (P<0.05): in the first birth milk yield, the fourth milk yield, one-year-old wool length and two-year-old thickness, the individuals with genotype GA were significant higher than genotype GG (P<0.05). It implied that Ex3 locus had significant effect on milk performance (milk yield) and wool performance (wool length and thickness).4.3 Associations of genetic variations of Ex4 locus with economic traits in goatThe mutation (EX4₅3delA) caused the six populations to two genotypes (AA and AD genotype). The genotype AA and allele A were predominant in the analyzed populations whose PIC values varied from 0.000 to 0.2800. Distribution of genotypes and alleles were founded to be significantly associated with characteristics of breeds (P<0.05, P<0.01), as well as goat types (diary type, meat type and wool type).4.4 Associations of genetic variations of Ex5 locus with economic traits in goatThe mutations (EX5₁4G>A, EX5₃4G>A,EX5₅9G>A) cause to analyzed populations to three genotypes (GG, GA and AA). The distributions of genotype GG and allele G were predominant. It was significantly found that the distribution of genotypes and alleles associate with the breed factor and the goat use types (dairy type, meat type and wool types) (P<0.05 and P<0.01).The relationship between this locus and milk performance and wool performance were observed (P<0.05): the average milk yield of individuals with genotype GG were significant higher than that of genotype GA (P<0.05), while the average wool yield and three-year-old wool length of individuals with genotype GA were prior to that of genotype GG. Hence, Ex5 locus demonstrated significantly gene effect on milk performance (milk yield) and wool performance (wool yield and length). 4.5 Associations of AluI polymorphism of Ex6 locus with economic traits in goatThese mutations (EX6₅8T>C, EX6₁72T>C, 3'UTR+92G>A, 3'UTR+110T>C) resulted in the polymorphism of Ex6 locus. Among them, EX₁72T>C variation formed the AluI RFLP. Three genotypes (TT, TC and CC) were detected and genotype TT and allele T were predominant in the analyzed populations by AluI PCR-RFLP methods. The AluI polymorphism was found to be significantly associated with the breed factor and the goat use types (dairy type, meat type and wool type) (P<0.05 and P<0.01). The relationships between the locus and milk performance and birth weight were observed (P<0.05): the average milk yield of individuals with genotype TC were prior to that of genotype TT (P<0.05), while birth weight of individuals with genotype TT were prior to that of genotype TC. Hence, the AluI polymorphism of Ex6 locus demonstrated significantly genetic effects on milk performance (milk yield) and growth traits.4.6 Associations of the DdeI polymorphism of Ex6 locus with economic traits in goatEX₅8T>C variation formed the DdeI RFLP. Two genotypes (D1D1, D1D2) were detected and genotype D1D1 and allele D1 were predominant in the analyzed populations by DdeI PCR-RFLP methods. The DdeI polymorphism was found to be significantly associated with the breed factor and the goat use types (dairy use, meat use and wool use) (P<0.05 and P<0.01). The associations of this locus with litter size, milk yield and one-year-old weight were observed (P<0.05): the litter size of individuals with genotype D1D1 were prior to that of genotype D1D2 (P<0.05), as well as milk yield and one-year-old weight. Hence, the DdeI polymorphism of Ex6 locus affected significantly reproduction, milk performance and growth development.4.7 The interact effects of double loci of POU1F1 gene on economic traits in goatThe interact effects of Ex1 and Ex4 loci had significantly effects on the third milk yield (P<0.05) and the combinated genotype GG-AA (Ex1-Ex4) was the best. The interact effects of Ex3 and Ex4 loci had significantly effects on average milk yield (P<0.05) and the combinated genotype GG-AD (Ex3-Ex4) was the best. The interact effects of Ex3 and Ex5 loci had significantkly effects on average milk yield (P<0.05) and the combinated genotype GA-GG (Ex1-Ex5) was the best. The interact effects of Ex1 and Ex5 loci had significantly effects on heart girth (P<0.05) and the combinated genotype GT-GG (Ex1-Ex5) was prior to others. The interact effects of Ex1 and Ex3 loci had significantly effects on body size (P<0.05) and the combinated genotype GT-GA (Ex1-Ex3) was prior to others.4.8 The genetic effects of goat POU1F1 gene on economic traits in goat Goat POU1F1 gene had significant effects on reproduction traits (litter size) (P<0.05), such as the IVS1+10G>T in intron 1, EX₅8T>C in exon 6. This gene showed significantly effects on milk performance (e.g. milk yield), such as exon 3, 5 and 6, intron 3 and 4, and 3'UTR (P<0.05), as well as the combinated genotypes of Ex1, Ex3 and Ex5 loci. Moreover, it affected significantly the growth development, such as AluI polymorphism, DdeI polymorphism, and combinated genotypes among Ex1, Ex3 and Ex5 loci. It was also implied that genetic effects of POU1F1 gene on wool performance (milk yield, wool length and wool thickness), such as Ex3 and Ex5 loci.5 Associations of genetic variations of PRL gene and goat economic traitsThe PCR-SSCP and DNA sequencing methods were applied to scan the genetic variations of exon 1～5 of goat Prolatin (PRL) gene and the results showed polymorphisms in exon 4 and exon 5. The X76049:g.332T>A resulting in 111Met>Lys and X76049:g.492G>T resulting in 164Leu>Phe, caused the polymorphisms of exon 4. Moreover, the X76049:g.576C>A resulting in 176Pro>Thr cause the polymorphism of exon 5.In exon 4 locus, the pattern A was predominant in the 3 patterns. This locus associated with the breed factor and the goat use types (dairy use, meat use and wool use) (P<0.05 and P<0.01). The associations of the locus with milk yield and wool length were observed (P<0.05): the milk yield of individuals with pattern A were prior to that of pattern C (P<0.05), while the wool length of pattern B and C were significantly higher than pattern A. Hence, the locus affected significantly milk performance and wool performance.In exon 5 locus, genotype CC and allele C were predominant in the 3 genotypes and two alleles in analyzed populations whose PIC values varied from 0.113～0.280. This locus was significantly associated with seven breeds and the goat use types (dairy use, meat use and wool use) (P<0.05 and P<0.01).In the birth weight, one-year-old stature, one-year-old heart girth, and four-year-old weight, the individuals with genotype CA were prior to genotype CC (P<0.05), while the wool thickness of genotype CC was significantly higher than genotype CA (P<0.05), hence, the locus affected significantly growth development and wool performance.The interact effects of exon 4 and exon 5 loci had significant effects on two-year-old wool yield (P<0.05) and the combinated genotype B-CC (Exon 4-Exon 5) was the best. So, the goat PRL gene had significantly effect on milk performance, wool performance and growth development.6 Associations of polymorphisms of IGFBP-3 gene with goat economic traits In IGFBP-3 loci, EX2₅8C>T, EX2₆7C>G, IVS2+78A>G and IVS2+217G>A were found in the analyzed populations. Among them, HaeIII and XspI PCR-RFLP methods could determinate the variation of EX₆7C>G and IVS2+217G>A, respectively. Their distributions of genotype and alleles were significantly associated with breed factors and goat use types (P<0.05 and P<0.01).The associations of the HaeIII polymorphism with litter size and wool performance were observed (P<0.05): the litter size of individuals with genotype H1H1 were prior to that of genotype H1H2 (P<0.05); the two-year-old wool yield of individuals with genotype H1H1 were higher than that of genotype with H2H2, while the five-year-old wool yield of genotype H2H2 were higher than those of genotype H1H1. The correlations of XspI polymorphism with growth traits were observed (P<0.05): The one-year-old body size of genotype X1X1 was higher than that of X2X2, while the one-year-old heart-girth of genotype X1X2 was prior to other genotypes (P<0.05).7 The genetic variation of goat LALBA geneThe M63868:g.1897T>C resulting in p.L100P, produced the polymorphism of exon 3 of goat LALBA gene. This mutation could genotyped by MspI. In analyzed populations, two genotypes was observed and PIC values varied from 0.000 to 0.0461 which were significantly lower than Italian population (P<0.05). The LALBA gene of Chinese goat was charactrerized by poor polymorphism, but middle polymorphism for Italian populations. The average wool yield of genotype LALBA-A1/A2 was prior to that of genotype LALBA-A1/A1 (P<0.05).8 Genetic variation of goat CSN1S1 geneThe genotypes A1A1 and A1A0 were detected in analyzed populations in CSN1S1 locus. The genotype A1A1 was predominant in populations, as well as allele A1. The associations of the locus with litter size and one-year-old body size were observed (P<0.05): the first birth litter size and average litter size of individuals with genotype A1A0 were higher than those of genotype A1A1, as well as one-year-old body size. It implied that CSN1S1 gene had significantly effects on reproduction and growth traits (P<0.05).