Objective: The purpose of this study was to investigate the genetic effects of six keratin (KRT) genes on the wool traits of 418 Chinese Merino (Xinjiang type) (CMXT) individuals. Methods: To explore the effects and association of six KRT genes on sheep wool traits, The polymerase chain reaction-bas...
Objective: The purpose of this study was to investigate the genetic effects of six keratin (KRT) genes on the wool traits of 418 Chinese Merino (Xinjiang type) (CMXT) individuals. Methods: To explore the effects and association of six KRT genes on sheep wool traits, The polymerase chain reaction-based single-strand conformation polymorphism (PCR-SSCP), DNA sequencing, and the gene pyramiding effect methods were used. Results: We report 20 mutation sites (single-nucleotide polymorphisms) within the six KRT genes, in which twelve induced silent mutations; five induced missense mutations and resulted in $Ile{\rightarrow}Thr$, $Glu{\rightarrow}Asp$, $Gly{\rightarrow}Ala$, $Ala{\rightarrow}Ser$, $Se{\rightarrow}His$; two were nonsense mutations and one was a same-sense mutation. Association analysis showed that two genotypes of the KRT31 gene were significantly associated with fiber diameter (p<0.05); three genotypes of the KRT36 gene were significantly associated with wool fineness score and fiber diameter (p<0.05), three genotypes of the KRT38 gene were significantly associated with the number of crimps (p<0.05); and three genotypes of the KRT85 gene were significantly associated with wool crimps score, body size, and fiber diameter (p<0.05). Analysis of the gene pyramiding effect between the different genotypes of the gene loci KRT36, KRT38, and KRT85, each genotype in a gene locus was combined with all the genotypes of another two gene loci and formed the different three loci combinations, indicated a total of 26 types of possible combined genotypes in the analyzed population. Compared with the other combined genotypes, the combinations CC-GG-II, CC-HH-IJ, CC-HH-JJ, DD-HH-JJ, CC-GH-IJ, and CC-GH-JJ at gene loci KRT36, KRT38, and KRT85, respectively, had a greater effect on wool traits (p<0.05). Conclusion: Our results indicate that the mutation loci of KRT31, KRT36, KRT38, and KRT85 genes, as well as the combinations at gene loci KRT36, KRT38, and KRT85 in CMXT have significant effects on wool traits, suggesting that these genes are important candidate genes for wool traits, which will contribute to sheep breeding and provide a molecular basis for improved wool quality in sheep.
Objective: The purpose of this study was to investigate the genetic effects of six keratin (KRT) genes on the wool traits of 418 Chinese Merino (Xinjiang type) (CMXT) individuals. Methods: To explore the effects and association of six KRT genes on sheep wool traits, The polymerase chain reaction-based single-strand conformation polymorphism (PCR-SSCP), DNA sequencing, and the gene pyramiding effect methods were used. Results: We report 20 mutation sites (single-nucleotide polymorphisms) within the six KRT genes, in which twelve induced silent mutations; five induced missense mutations and resulted in $Ile{\rightarrow}Thr$, $Glu{\rightarrow}Asp$, $Gly{\rightarrow}Ala$, $Ala{\rightarrow}Ser$, $Se{\rightarrow}His$; two were nonsense mutations and one was a same-sense mutation. Association analysis showed that two genotypes of the KRT31 gene were significantly associated with fiber diameter (p<0.05); three genotypes of the KRT36 gene were significantly associated with wool fineness score and fiber diameter (p<0.05), three genotypes of the KRT38 gene were significantly associated with the number of crimps (p<0.05); and three genotypes of the KRT85 gene were significantly associated with wool crimps score, body size, and fiber diameter (p<0.05). Analysis of the gene pyramiding effect between the different genotypes of the gene loci KRT36, KRT38, and KRT85, each genotype in a gene locus was combined with all the genotypes of another two gene loci and formed the different three loci combinations, indicated a total of 26 types of possible combined genotypes in the analyzed population. Compared with the other combined genotypes, the combinations CC-GG-II, CC-HH-IJ, CC-HH-JJ, DD-HH-JJ, CC-GH-IJ, and CC-GH-JJ at gene loci KRT36, KRT38, and KRT85, respectively, had a greater effect on wool traits (p<0.05). Conclusion: Our results indicate that the mutation loci of KRT31, KRT36, KRT38, and KRT85 genes, as well as the combinations at gene loci KRT36, KRT38, and KRT85 in CMXT have significant effects on wool traits, suggesting that these genes are important candidate genes for wool traits, which will contribute to sheep breeding and provide a molecular basis for improved wool quality in sheep.
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문제 정의
The results revealed 20 mutation sites, IVS2+50-52insG, EX3_62delA, IVS2+85G/A, IVS2+95C/T, EX7_113G/A, EX7_117C/A, EX7 _118T/A, EX7_119T/C, EX7_120G/A, EX7_121delC, EX3_ 49delA, and EX7_111G/C, which showed silent mutations; among all mutations, EX7_93T/C, EX7_97A/C, EX7_112A/T, EX7_126G/T, and EX3_63T/A, showed missense mutations resulting in Ile→Thr, Glu→Asp, Gly→Ala, Ala→Ser, and Se→His; nonsense mutations EX3_83A/T and EX3_210G/A; and a same-sense mutation EX3_354G/A. An association between different genotypes of KRT31, KRT36, KRT38, and KRT85 and wool traits in CMXT was established in this study. These data strongly suggest that gene polymorphisms may represent genetic markers that could be used for the breeding of new sheep breeds.
가설 설정
1) Data within parentheses are the number of individuals with different genotypes.
제안 방법
In this study, PCR-SSCP and DNA sequencing methods were used to identify polymorphisms in the sheep KRT27, KRT31, KRT36, KRT38, KRT81, and KRT85 genes. Only one SSCP genotype was identified in the P1, P2, P3, and P4 fragments of KRT27, P6 and P7 fragments of KRT36, P9 fragment of KRT38, P12, P13, P14, and P15 fragments of KRT81, and P17 fragment of KRT85.
On the basis of the sequences of the KRT27, KRT31, KRT36, KRT38, KRT81, and KRT85 genes obtained from the National Center for Biotechnology Information GenBank database (GenBank accession numbers AC_000176[521074], AC_ 000176[539597], AC_000176[520668], NC_00176[515000], AC_000162[540204], AC_000162[528459], respectively), 17 pairs of PCR primers (P1−P17, Table 1) were designed to amplify different PCR products including exon3, exon8, intervening sequence 2 (IVS2), IVS5 of KRT27; IVS2 of KRT31; exon2, exon3 of KRT36; exon3, exon7, IVS2 of KRT38; exon1, exon2, IVS1, IVS3 of KRT81; and exon3, IVS2 of KRT85.
The main objective of this study was to analyze the possible polymorphisms in 17 primer fragments of six candidate genes of the keratin family, including KRT27 (exons 3 and 8, introns 2 and 5), KRT31 (intron 2), KRT36 (exons 1, 2, and 3), KRT38 (exon 3, introns 2 and 7), KRT81 (exons 1 and 2, introns 1 and 3), KRT85 (exon 3, intron 2) in CMXT using the polymerase chain reaction-based single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing methods. Association between the identified polymorphism sites, as well as the combined genotypes, with wool traits (staple length, wool crimps score, number of crimps, wool fineness, body size, live weight after shearing, greasy weight, fiber diameter, and coefficient of variation) were tested by least-square analysis.
대상 데이터
Genomic DNA samples were obtained from 418 healthy CMXT, which were reared at Gonaisi Fine Wool Sheep Breeding Farm, Xinyuan county, (Latitude 43°03′–43°41′N, Longitude 82°28′– 84°56′E), Xinjiang Province, China.
데이터처리
The polymorphic information content (PIC) was calculated using the method described by [21]. A chi-square test was applied to identify statistical significance, which was performed using SAS 8.1 software (SAS Inc., Cary, NC, USA) [22]. Associations between different genotypes and wool traits were analyzed by analysis of variance (SAS 8.
The main objective of this study was to analyze the possible polymorphisms in 17 primer fragments of six candidate genes of the keratin family, including KRT27 (exons 3 and 8, introns 2 and 5), KRT31 (intron 2), KRT36 (exons 1, 2, and 3), KRT38 (exon 3, introns 2 and 7), KRT81 (exons 1 and 2, introns 1 and 3), KRT85 (exon 3, intron 2) in CMXT using the polymerase chain reaction-based single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing methods. Association between the identified polymorphism sites, as well as the combined genotypes, with wool traits (staple length, wool crimps score, number of crimps, wool fineness, body size, live weight after shearing, greasy weight, fiber diameter, and coefficient of variation) were tested by least-square analysis. These findings provide a scientific basis for the improvement of wool traits through marker-assisted selection (MAS).
이론/모형
Based on the genotypes of 17 KRT fragments in the analyzed population, genotype frequencies, allele frequencies, and Hardy−Weinberg equilibrium were calculated.
Based on the genotypes of 17 KRT fragments in the analyzed population, genotype frequencies, allele frequencies, and Hardy−Weinberg equilibrium were calculated. The population genetic indices, such as He (gene heterozygosity), Ho (gene homozygosity; Ho+He = 1), and Ne (effective allele numbers; reciprocal of homozygosity) were calculated using the PopGene software (version 3.2, University of Alberta, Edmonton, AB, Canada). The polymorphic information content (PIC) was calculated using the method described by [21].
성능/효과
Combined genotypes indeed had more profound impacts than the individual genotypes [25,26]. Based on the analysis of the combined genotypes of KRT36, KRT38, and KRT85, a total of 26 superior combined genotypes were found in the analyzed population, among which the combined genotype DD-GG-II was dominant. Individuals with the combined genotypes CC-GG-II had longer staple length, those with the combined genotypes CC-HH-IJ had a higher wool crimps score and number of crimps, individuals with combined genotypes CC-HH-JJ had higher wool fineness, individuals with combined genotypes DD-HH-JJ had higher body size, individuals with combined genotypes CCGH-IJ had greater live weight after shearing, individuals with combined genotypes CC-GH-JJ had higher greasy weight, fiber diameter, and coefficient of variation, Therefore, these combined genotypes represent the optimal combinations, suggesting that KRT36, KRT38, and KRT85 should be used as candidate genes for sheep breeding.
In conclusion, 20 mutation sites extended the characterization of genetic variation in the KRT31, KRT36, KRT38, and KRT85 genes. Additionally, we demonstrated significant associations between the sheep KRT31, KRT36, KRT38, and KRT85 genes and wool traits.
In the P16 fragment of KRT85 gene, three SSCP banding patterns were found, named II, IJ, and JJ, respectively, Figure 1E. In order to better characterize genetic variation of the P16 fragment within the sheep KRT85 gene, the amplified polymorphic DNA fragments were sequenced and three mutations (a silent mutation EX3_49delA, a nonsense mutation EX3_ 210G/A, and a same-sense mutation EX3_354G/A), were detected (Figure 2E). The frequencies of alleles I and J were 0.
Additionally, we demonstrated significant associations between the sheep KRT31, KRT36, KRT38, and KRT85 genes and wool traits. Moreover, based on the findings of combined genotype analysis, the CC-GG-II, CC-HH-IJ, CC-HH-JJ, DD-HH-JJ, CC-GH-IJ, and CC-GH-JJ combined genotypes for CMXT could be used as molecular markers to identify superior wool traits, and KRT31, KRT36, KRT38, and KRT85 are potential candidate genes for wool traits. However, the present study presents preliminary results and further investigations are essential.
In this study, PCR-SSCP and DNA sequencing methods were used to identify polymorphisms in the sheep KRT27, KRT31, KRT36, KRT38, KRT81, and KRT85 genes. Only one SSCP genotype was identified in the P1, P2, P3, and P4 fragments of KRT27, P6 and P7 fragments of KRT36, P9 fragment of KRT38, P12, P13, P14, and P15 fragments of KRT81, and P17 fragment of KRT85.
These data strongly suggest that gene polymorphisms may represent genetic markers that could be used for the breeding of new sheep breeds. Results of the combined genotypes analysis indicated that a group of six combined genotypes for CMXT polymorphisms led to the appearance of better wool traits, including staple length in the CC-GG-II genotype, wool crimps score and number of crimps in the CC-HH-IJ genotype, wool fineness in the CC-HH-JJ genotype, body size in the DD-HH-JJ genotype, live weight after shearing in the CC-GH-IJ genotype, and greasy weight, fiber diameter, and coefficient of variation in the CC-GH-JJ genotype.
The results revealed 20 mutation sites, IVS2+50-52insG, EX3_62delA, IVS2+85G/A, IVS2+95C/T, EX7_113G/A, EX7_117C/A, EX7 _118T/A, EX7_119T/C, EX7_120G/A, EX7_121delC, EX3_ 49delA, and EX7_111G/C, which showed silent mutations; among all mutations, EX7_93T/C, EX7_97A/C, EX7_112A/T, EX7_126G/T, and EX3_63T/A, showed missense mutations resulting in Ile→Thr, Glu→Asp, Gly→Ala, Ala→Ser, and Se→His; nonsense mutations EX3_83A/T and EX3_210G/A; and a same-sense mutation EX3_354G/A.
1 Itenge TO Hickford J Forrest R Mckenzie GW Frampton CM Improving the quality of wool through the use of gene markers S Afr J Anim Sci 2010 39 219 23
2 Yu Z Wildermoth JE Wallace OAM Annotation of sheep keratin intermediate filament genes and their patterns of expression Exp Dermatol 2011 20 582 8 21554405
4 Coulombe PA Omary MB ‘Hard’ and ‘soft’ principles defining the Structure, function and regulation of keratin intermediate filaments Curr Opin Cell Biol 2002 14 110 22 11792552
12 Bawden CS Sivaprasad AV Verma PJ Walker SK Rogers GE Expression of bacterial cysteine biosynthesis genes in transgenic mice and sheep: toward a new in vivo amino acid biosynthesis pathway and improved wool growth Transgenic Res 1995 4 87 104 7704055
13 Powell B Crocker L Rogers G Hair follicle differentiation: expression, structure and evolutionary conservation of the hair type II keratin intermediate filament gene family Development 1992 114 417 33 1375545
14 Powell BC Beltrame JS Characterization of a hair (wool) keratin intermediate filament gene domain J Invest Dermatol 1994 102 171 7 7508962
16 Yu Z Gordon SW Nixon AJ Expression patterns of keratin intermediate filament and keratin associated protein genes in wool follicles Differentiation 2009 77 307 16 19272529
18 Langbein L Rogers MA Winter H The catalog of human hair keratin. I: expression of the nine type I members in the hair follicle J Biol Chem 1999 274 19874 84 10391933
19 Di J Wang Q Li Y LAZHTI The keratinis genes and its expression in the wool follicle Grass-feeding Livest 2004 17 19
20 Sun HS Anderson LL Yu TP Neonatal Meishan pigs show POU1F1 genotype effects on plasma GH and PRL concentration Anim Reprod Sci 2002 69 223 37 11812632
21 Botstein D White RL Skolnick M Darvis RW Construction of a genetic linkage map in man using restriction fragment length polymorphisms Am J Hum Genet 1980 32 314 31 6247908
26 Rodriguez S Eiriksdottir G Gaunt TR IGF2BP1 , IGF2BP2 and IGF2BP3 genotype, haplotype and genetic model studies in metabolic syndrome traits and diabetes Growth Horm IGF Res 2010 20 310 8 20627640
27 Li-Juan LI Song DR Han FX Cloning and sequence analysis of goat KIFI gene China Anim Husb Vet Med 2010 37 91 3
28 Wu X Tian KC Liu WJ Construction of a long-lenth cDNA library for Xinji Fine-wool Sheep skin tissue by use of SMART technique J Northwest A & F Univ 2010 38 46 50
29 Tian YZ Jiang DI Wei-Wei WU Expression difference of keratin 26 gene of skin tissues in superfine and fine Merino sheep China Anim Husb Vet Med 2012 39 31 7
30 Juanjuan YU Liu J Zhao J Gene chip analysis of expression pattern of type i inner root sheath (IRS) keratin in AoHan wool sheep Agric Sci Technol 2012 13 1171 4
31 Wang LP Yang BH Yue YJ Analysis on genetic polymorphism of KRT35 gene sequence of Gansu alpine fine-wool sheep Guizhou Agric Sci 2010 38 142 4
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