• Asian-Australasian Journal of Animal Sciences
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• 제21권4호
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• pp.465-470
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• 2008

This study describes genetic variability in the BoLA-DRB3 gene in Iranian native cattle (Bos Indicus and Taurus) and relationships between these breeds. This is the first study of genetic polymorphism of the BoLA-DRB3 gene in Iranian native cattle. We examined exon 2 of the major histocompatibility complex (MHC) class II DRB3 gene from 203 individuals in four populations of Iranian native cattle (52 Sarabi, 52 Najdi, 49 Sistani, 50 Golpayegani cattle) using the hemi-nested PCR-RFLP method. We identified the 36 previously reported alleles and one novel pattern (*eac). Analysis of the frequencies of the various BoLA-DRB3.2 alleles in each breed indicated that DRB3.2*52 in Sarabi cattle (23%), DRB3.2 *14 and *24 alleles in Najdi cattle (13%), DRB3.2 *8 allele in Sistani cattle (22%) and DRB3.2*16 allele in Golpayegani cattle (14%), were the most frequent alleles. Allelic frequencies ranged from 1 to 23% among the 36 alleles and there were some alleles that were found only in Iranian cattle. Effective number of alleles in the four breeds was estimated to be 7.86, 11.68, 7.08 and 3.37 in Sarabi, Najdi, Sistani and Golpayegani, respectively. Observed heterozygosities were the highest in Sarabi (94%) and Najdi (94%). A population tree based on the frequency of BoLA-DRB3.2 alleles in each breed suggested that Najdi, Sarabi and Golpayegani cattle clustered together and Najdi and Sarabi were the closest breeds. Sistani cattle differed more from these three breeds. These new data suggest that allele frequencies differ between Iranian cattle breeds.

• Asian-Australasian Journal of Animal Sciences
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• 제18권12호
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• pp.1691-1695
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• 2005

The bovine lymphocyte antigen (BoLA)-DRB3 gene encodes cell surface glycoproteins that initiate immune responses by presenting processed antigenic peptides to CD4 T helper cells. DRB3 is the most polymorphic bovine MHC class II gene which encodes the peptide-binding groove. Since different alleles favour the binding of different peptides, DRB3 has been extensively evaluated as a candidate marker for associations with various bovine diseases and immunological traits. For that reason, the genetic diversity of the bovine class II DRB3 locus was investigated by polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFLP). This study describes genetic variability in the BoLA-DRB3 in Iranian Golpayegani Cattle. Iranian Golpayegani Cows (n = 50) were genotyped for bovine lymphocyte antigen (BoLA)-DRB3.2 allele by polymerase chain reaction and restriction fragment length polymorphism method. Bovine DNA was isolated from aliquots of whole blood. A two-step polymerase chain reaction followed by digestion with restriction endonucleases RsaI, HaeIII and BstYI was conducted on the DNA from Iranian Golpayegani Cattle. In the Iranian Golpayegani herd studied, we identified 19 alleles.DRB3.2${\times}$16 had the highest allelic frequency (14%), followed by DRB3.2${\times}$7 (11%). Six alleles (DRB3.2${\times}$25, ${\times}$24, ${\times}$22, ${\times}$20, ${\times}$15, ${\times}$3) had frequencies = 2%. Although additional studies are required to confirm the present findings, our results indicate that exon 2 of the BoLA-DRB3 gene is highly polymorphic in Iranian Golpayegani Cattle.

• 농업과학연구
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• 제25권1호
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• pp.79-88
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• 1998

본 연구는 축협중앙회 한우개량부에서 사육중인 한우의 혈액으로부터 genomic DNA를 추출하여 한우의 면역체계에 중요한 역할을 담당하고 있는 BoLA DRB3 exon2 유전자를 PCR기법을 이용하여 증폭하고 친자확인을 위한 이들 대립유전자들의 염기서열을 분석하여 한우의 효율적인 육종에 분자유전수준에서의 접목을 위한 기초자료를 얻고자 실시하였던바, 얻어진 결과를 요약하면 다음과 같다. 1 한우의 혈액에서 추출된 genomic DNA를 1.5% agarose gel에서 전기영동한 결과, 12.2kb이상의 단일밴드로 나타나, genomic DNA는 아주 잘 분리된 것으로 판단되었으며, 한편 genomic DNA로부터 PCR기법을 이용하여 BoLA DRB3 exon2 유전자를 증폭한 결과 284kb의 단편이 증폭 되었음을 확인하였다. 2. PCR 증폭산물을 pCR 2.1 vector를 사용하여 cloning 하고, 균주에 형질전환을 시켜 재조합 plasmid를 추출한 후, EcoR 1으로 처리한 결과 300bp 단편이 확인되어 증폭되어진 BoLA DRB3 exon2 유전자가 vector 내에 잘 삽입되었음을 확인하였다. 3. 부와 모의 BoLA DRB3 exon2 대립유전자의 염기서열을 분석한 결과 염기서열의 상동성은 부의 대립유전자간에는 82.0% 이었고, 모의 대립유전자간에는 90.1%이었다. 4. 친자를 확인하기 위해 부와 모 그리고 자손간의 가계에 대한 BoLA DRB3 exon2 대립유전자의 염기서열을 분석한 결과 Mendel 의 법칙에 따라 유전된다는 사실을 확인 할 수 있었다. 5. 이상의 결과를 종합하여보면 한우의 BoLA DRB3 exon2 유전자의 부와 모, 그리고 자손의 대립유전자에 대한 염기서열 수준에서의 친자확인이 가능하였으며, 이들 연구결과는 축우의 유전적 능력개량에 중요한 분자유전학적 기초 자료가 될 것으로 판단되었다.

• Journal of Animal Science and Technology
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• 제63권2호
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• pp.248-261
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• 2021

Attributable to their major function in pathogen recognition, the use of bovine leukocyte antigens (BoLA) as disease markers in immunological traits in cattle is well established. However, limited report exists on polymorphism of the BoLA gene in zebu cattle breeds by high resolution typing methods. Thus, we used a polymerase chain reaction sequence-based typing (PCR-SBT) method to sequence exon 2 of the BoLA class II DRB3 gene from 100 animals (Boran, n = 13; Sheko, n = 20; Fogera, n = 16; Horro, n = 19), Hanwoo cattle (n = 18) and Bangladesh Red Chittagong zebu (n = 14). Out of the 59 detected alleles, 43 were already deposited under the Immuno Polymorphism Database for major histocompatibility complex (IPD-MHC) while 16 were unique to this study. Assessment of the level of genetic variability at the population and sequence levels with genetic distance in the breeds considered in this study showed that Zebu breeds had a gene diversity score greater than 0.752, nucleotide diversity score greater than 0.152, and mean number of pairwise differences higher than 14, being very comparable to those investigated for other cattle breeds. Regarding neutrality tests analyzed, we investigated that all the breeds except Hanwoo had an excess number of alleles and could be expected from a recent population expansion or genetic hitchhiking. Howbeit, the observed heterozygosity was not significantly (p < 0.05) higher than the expected heterozygosity. The Hardy Weinberg equilibrium (HWE) analysis revealed non-significant excess of heterozygote animals, indicative of plausible over-dominant selection. The pairwise FST values suggested a low genetic variation among all the breeds (FST = 0.056; p < 0.05), besides the rooting from the evolutionary or domestication history of the cattle. No detached clade was observed in the evolutionary divergence study of the BoLA-DRB3 gene, inferred from the phylogenetic tree based on the maximum likelihood model. The investigation herein indicated the clear differences in BoLA-DRB3 gene variability between African and Asian cattle breeds.

• Asian-Australasian Journal of Animal Sciences
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• 제20권12호
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• pp.1791-1797
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• 2007

A study was conducted with 70 Hanwoo (Korean cattle) for genotyping bovine leukocyte antigen (BoLA)-DRB3.2 gene by using the polymerase chain reaction (PCR) and sequence based typing (SBT). Two-step PCR was carried out for amplifying a 284 bp fragment of the target gene and the PCR products were digested with three restriction enzymes namely RsaI, BstYI and HaeIII. Seventeen alleles were detected with frequencies ranging from 1.43 to 18.57% and one (x'aa) of these alleles was identified as a new allele that has not been reported before. The frequency of the new x'aa allele identified in this breed was 12.86%. In addition, the seven most frequently observed alleles (DRB3.2 *10, *15, *16, *26, *27, *54 and x'aa) accounted for 74.28% of the alleles in this population. The phylogenetic tree showed that the BoLA-DRB3.2 allele sequences of Hanwoo were shared with other Bos taurus breeds and no specific clade for Hanwoo was identified. It indicates high heterogeneity of the BoLA-DRB3 gene in this population and may give some ideas for breeding animals having better disease resistance.

• 농업과학연구
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• 제38권1호
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• pp.51-63
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• 2011

BoLA (bovine leukocyte antigens) have been known as gene complex related with bovine diseases and immunological traits. This study was conducted to find out the characteristics of BoLA-DRB3 gene related to mastitis and BL(bovine leukocyte) from 280 cattle [193 animals of Holstein cattle and 87 animals of Hanwoo]. As a result, five PCR-RFLP types (b, d, e, f and g) using HaeIII restriction enzyme, three BstYI restriction patterns (b, d and e) and eight RsaI restriction types(b, d, f, I, j, n, o and w) were identified. Moreover, we identified new d' type ($197{\rightarrow}175$/22), having one more cutting site by BstYI enzyme than d type allele and n' type ($180{\rightarrow}169$/11) having one more cutting site by RsaI enzyme than n allele was additionally identified. Next, we identified 53 SNPs in BoLA-DRB3 exon2 from 280 cattle. SNP frequency and heterozygosity of Holstein and Hanwoo were investigated in all the SNP genotype. These results might be based on research for identifying marker associated with bovine diseases.

• Asian-Australasian Journal of Animal Sciences
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• 제19권6호
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• pp.775-778
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• 2006

Sarabi cows (n = 136) from the Sarabi Breeding Station were genotyped at bovine lymphocyte antigen (BoLA)-DRB3.2 locus by a genotyping system that used the polymerase chain reaction and restriction fragment length polymorphism. Genomic DNA was extracted from whole blood samples. A two-step polymerase chain reaction was carried out in order to amplify a 284 base-pair fragment of target gene. Nested-PCR products were digested with three restriction endonuclease enzymes RsaI, BstYI and HaeIII. Digested fragments were analyzed by polyacrylamide gel electrophoresis. Twenty-six BoLA-DRB3.2 alleles were identified with frequencies ranging from 0.4 to 15.1%. Six new allele types observed in this study have not been reported previously. Identified alleles include: BoLA-DRB3.$2^*1$, $^*2$, $^*4$, $^*6$, $^*8$, $^*12$, $^*13$, $^*14$, $^*15$, $^*16$, $^*17$, $^*23$, $^*24$, $^*25$, $^*28$, $^*32$, $^*34$, $^*35$, $^*36$, $^*37$, $^*42$, $^*46$, $^*51$, $^*kba$, $^*laa$ and $^*vaa$. Their frequencies were found to be 0.4, 0.4, 0.7, 11.4, 1.1, 1.8, 2.9, 2.2, 4.4, 9.6, 1.1, 13.6, 0.4, 0.4, 1.1, 0.7, 0.4, 6.2, 2.2, 3.7, 1.1, 7.7, 1.5, 15.1, 2.6 and 7.3% respectively. The six most frequent alleles (DRB3.2 $^*6$, $^*16$, $^*23$, $^*46$, $^*kba$ and $^*vaa$) accounted for 64.7% of the alleles in the population of this herd. Numerous studies on this locus, covering different breeds, has revealed the existence of various alleles in this locus, and new investigations have introduced novel alleles. With respect to the high number of the observed alleles in this survey and the novelty of some alleles with no previous record of reporting, it is plausible to conclude that the BoLA-DRB3.2 locus is highly polymorphic in Iranian native Sarabi cows.

• Asian-Australasian Journal of Animal Sciences
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• 제22권11호
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• pp.1487-1494
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• 2009

The major histocompatibility complex (MHC) plays well-defined roles in eliciting immune responses and combating infectious diseases. The major histocompatibility complex of cattle is referred to as BoLA (Bovine Lymphocyte Antigen). This genetic system is among the most polymorphic. In the present study, polymorphism of the BoLA- DRB3.2 locus in three Bos indicus breeds viz., Sahiwal, Rathi and Hariana was studied by polymerase chain reaction restriction fragment length polymorphism technique using the enzymes RsaI, Bst Y1 and Hae III. Both Sahiwal and Rathi are good Indian dairy breeds and survive under tough tropical conditions, while Hariana is a prominent dual-purpose breed reared both as a dairy animal and for bullock production. A total of 30 different BoLADRB3.2 alleles were observed to be present in the 3 Bos indicus breeds. Certain alleles were common amongst the three breeds while there were others that were unique to each breed. Allelic distribution amongst the three breeds showed that each breed had a unique allelic distribution pattern that was different from each other and also different from the earlier breeds studied so far for the existence of allelic variation at this locus. A dendogram was constructed based on the frequencies of the BoLA-DRB3 alleles using the UPGMA method. The Rathi and Hariana animals were genetically the most apart. The Hariana animals clustered on a different branch from the other two breeds viz. the Rathi and the Sahiwal. The smallest genetic distances for the DRB3 alleles were those between Sahiwal and Rathi (0.5461) while genetic distance between Hariana and Sahiwal was 0.6123. A comparison of the allelic frequencies of the BoLADRB3.2 locus in these 3 breeds viz. Sahiwal, Hariana and Rathi with the allelic frequencies present in the previously characterized Bos indicus Kankrej breed, which is a dual purpose breed reared both as a draught and a dairy animal, showed that the Bos indicus Sahiwal and Rathi breeds clustered into one group while the Hariana and Kankrej breeds formed another group. The Rathi and Sahiwal showed the least genetic distance of 0.5461 amongst the breeds whereas the Rathi and Kankrej, with a Nei''s genetic distance of 1.1622, were genetically the most distant apart.

• 한국동물학회:학술대회논문집
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• 한국동물학회 1997년도 한국생물과학협회 학술발표대회
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• pp.292.2-292
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• 1997

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