한우 개체식별 및 친자감정을 위한 새로운 초위성체 마커 개발 = Development of a New Microsatellite Markers for Individual Identification and Paternity Evaluation in Hanwoo

다국어 초록 (Multilingual Abstract)

The generally used eleven Di-nucleotide repeat microsatellite markers cause ‘stutter', which makes it difficult to estimate the accurate allele size. This study was conducted to solve this problem and to propose new thirteen microsatellite markers (BTRC6_01, BTRC19_02, BTRC11_03, BTRC16_05, BTRC9_07, BPC19_08, BTEC17_09, BPC21_10, BTEC4_11, BPC7_12, BPC1_13, BHXC29_14, BPC1_15) composed of tri-, tetra-, penta-, and hexa-nucleotide repeat microsatellite loci as genetic parameters for Hanwoo discrimination and genetic diversity analysis. After testing on 1530 cattle with 13 tested microsatellite loci, a total of 61 alleles were detected and the mean number of alleles per locus was 4.69. The Polymorphism Information Contents (PIC) ranged from 0.25(BTRC9_07) to 0.59(BTEC17_09). Since BHXC29_14, BPC1_13, BTEC17_09, BTRC16_05, BTRC19_02, and BTRC6_01 were highly informative(PIC>0.5) and the rest of the loci were reasonably informative (PIC>0.25), the thirteen loci are considered to have enough polymorphism for bovine identification.
Heterozygosity and FIS (inbreeding coefficient) value in all cattle population of 2 local brand Hanwoo (Yeongam Hanwoo;YH and Jangheung;JH) and 7 breeds of European cattle (Brown Swiss;BS, Limousin;LM, Angus;AG, Simmental;SM, Hereford;HF, Charolais;CH and Holstein;HT) were calculated to identify genetic diversity and characteristics of Hanwoo. The expected heterozygosity of Hanwoo was 0.532(YH) and 0.545(JH), compared to 0.451(BS)~0.605(AG). Phylogenetic analysis among Hanwoo and 7 breeds of European cattle was conducted by estimating Nei's genetic distance based on specific allele frequencies. Among other European breeds, SM showed the closest genetic distance (0.1848) to Hanwoo.

국문 초록 (Abstract)

본 연구는 한우 개체식별 및 친자감별에 있어 기존의 di-nucleotide repeat microsatellite marker 사용 시 발생했던 stutter로 인한 대립유전자판별 오류 등의 문제들을 극복하고, 분석결과의 신뢰도와 정확도를 높이기 위해 tri-, tetra-, penta-, hexa-nucleotide repeat microsatellite 좌위들로이루어진 새로운 개체식별 마커 13 종(BTRC6_01, BTRC19_02, BTRC11_03, BTRC16_05, BTRC9_07, BPC19_08, BTEC17_09, BPC21_10, BTEC4_11, BPC7_12, BPC1_13, BHXC29_14, BPC1_15)을 개발하였다. 선발된 13개의 좌위를 가지고 소 1,530두에 microsatellite typing을실시한 결과, 총 61개에 대립유전자가 발견되었으며, 좌위별로 평균 4.69개의 대립유전자를 가지는 것으로 확인되었다. 마커의 다형성과 정보력의척도인 PIC (Polymorphism Information Contents)값은 0.25(BTRC9_07)~0.59(BTEC17_09)로 나타났으며 BHXC29_14, BPC1_13, BTEC17_09, BTRC16_05, BTRC19_02, BTRC6_01 좌위들은 PIC 0.5 이상 그리고 나머지 좌위들 모두 PIC 0.25 이상의 값을 가지는 것으로 확인되어마커로서 다형성이 있음이 검증되었다. 개발한 마커를 활용하여 한우(영암, 장흥)와 유럽우 7종 (Brown Swiss, Limousin, Angus, Simmental, Hereford, Charolais, Holstein)의 유전적 특성을 분석하였으며, 총 9개 집단의 Heterozygosity와 FIS (inbreeding coefficient) 값을 측정하였다.
기대이형접합율은 0.451(BS)~0.605(AG) 범위 내로, 한우는 0.532(영암), 0.545(장흥)의 값을 가지는 것으로 나타났다. 한우(영암, 한우)와 7종의유럽우들 간의 유연관계 분석은 특정 대립유전자 빈도를 근거로 한 유전적 거리의 추정으로 이루어졌다. 한우집단과 Simmental 간의 유전적거리(0.1848)가 가장 가깝고 비교적으로 Brown Swiss와의 유전적 분포(0.3352)가 가장 먼 것으로 나타났으며, 계통발생학적으로 유전적 분화양상을 확인함으로써 본 마커의 한우 유전적 다양성 및 유연관계 분석에 활용 가능성을 제시하였다.

참고문헌 (Reference)

1 Saitou N, "The neighbor-joining method: A new method for reconstruction phylogenetic tree" 4 : 406-425, 1987

2 Benson G, "Tandem repeats finder : A program to analyze DNA sequences" 27 : 573-580, 1999

3 Van de Goor LHP, "Population studies of 16 bovine STR loci for forensic purposes" 125 : 111-119, 2009

4 Bacher JW, "Penta Nucleotide repeats: Highly polymorphic genetic markers displaying minimal stutter" Promega Corporation 1998

5 Chamber GK, "Microsatellites: Consensus and controversy" 126 : 455-476, 2000

6 Park SDE, "Microsatellite toolkit for MS Excel 97 or 2000"

7 이기환, "Microsatellite loci 분석을 통한 한우 지역 브랜드간 유전적 다양성의 비교" 한국축산학회 50 (50): 167-176, 2008

8 윤두학, "Microsatellite loci 분석에 의한 한우와 타 품종간의 유전적 유연관계" 한국축산학회 47 (47): 341-354, 2005

9 Chung HY, "Isolation and characterization of the bovine microsatellite loci" 44 : 527-541, 2006

10 Tautz D, "Hypervariability of simple sequences as a general source for polymorphic DNA marker" 17 : 6463-6471, 1989

11 Takezaki PJ, "Genetic distance and reconstruction of phylogenetic tree from microsatellite DNA" 144 : 389-399, 1996

12 Delgado JV, "Genetic characterization of Latin-American Creole cattle using microsatellite markers" 43 : 2-10, 2011

13 심용택, "Forensic Characterization of Four New Bovine Tri-nucleotide Microsatellite Markers in Korean Cattle(Hanwoo)" 한국축산학회 55 (55): 87-93, 2013

14 Lee SH, "Establishment of microsatellite marker set for the individual identification its application in the Hanwoo industry" GyengSang National University 2007

15 Du-Hak Yoon, "Establishment of an Individual Identification System Based on Microsatellite Polymorphisms in Korean Cattle (Hanwoo)" 아세아·태평양축산학회 18 (18): 762-766, 2005

16 Ota T, "DISPAN: Genetic distance and phylogenetic analysis" Pennsylvania State University 1993

17 Botstein D, "Construction of a genetic linkage map in man using restriction fragment length polymorphisms" 32 : 314-331, 1980

18 Fernandez ME, "Comparison of the effectiveness of microsatellites and SNP panels for genetic identification, traceability and assessment of parentage in an inbred Angus herd" 36 (36): 185-191, 2013

19 Nei M, "Accuracy of estimated phylogenetic trees from molecular data" 19 : 153-170, 1983

20 Barker JSF, "A global protocol for determining genetic distances among domestic livestock breeds" 21 : 501-508, 1994

21 Barendse W, "A genetic map of DNA loci bovine chromosome 1" 18 : 602-608, 1993