BoLA-DRB3 Gene Research Articles

Prevalence of bovine dermatophilosis and disease-associated alleles in zebu Goudali cattle and their Italian Simmental crosses ranching in the western highland plateau savannah of Cameroon.

Abundance of native pastures makes Cameroon's western highland savannah (WHS) a hotspot for low-input beef-type cattle. Dumbo Ranch is central to cattle seed stock multiplication in WHS and holds that Dermatophilus congolensis infection undermines production. The bovine BoLA-DRB3 has been variously demonstrated as the principal gene of the major histocompatibility locus associated with immunity and resistance to dermatophilosis in cattle. We studied the profile of dermatophilosis prevalence in zebu Goudali (G) and its Simmental composite, SimGoud (SG), at Dumbo Ranch and determined the distribution of a dermatophilosis-associated susceptibility allele of the BoLA-DRB3 gene by allele-specific polymerase chain reaction (PCR). We recorded a 42% prevalence of dermatophilosis in the studied cohort (337 animals). Dermatophilosis was more common in older cattle than in cattle ≤36months (p ≤ 0.05). G was more affected compared to SG, because of the prevalence of the disease in the oldest animals and the age distribution of the experimental subjects. No susceptible homozygote was observed. About 85 and 15% of the cohort carried the homozygous resistant and heterozygous condition, respectively. This genotype distribution was not affected by cattle type. The study confirms the presence of dermatophilosis among G and SG cattle in WHS. However, there was no correlation between the presence of the disease-associated susceptible allele considered and clinical manifestation. Screening for this dermatophilosis resistance-associated allele of BoLA-DRB3 gene appeared not useful for selection of G and SG in WHS.

Haplotype determination of the upstream regulatory region and the second exon of the BoLA‐DRB3 gene in Holstein cattle

Polymorphisms of the BoLA-DRB3 gene are located primarily in the second exon [antigen binding site (ABS)] and, to a lesser extent, in the upstream regulatory region (URR). It can be hypothesised that exon 2 and the URR are under different types of natural selection. The aim of this work was to determine the URR-exon 2 haplotypes; 34 Holstein samples were genotyped by direct sequencing. A total of 7 URR alleles and 23 exon 2 alleles were detected, and 3 of the URR alleles were novel. Our results may suggest that no relationship exists between the URR and exon 2 of the BoLA-DRB3 gene (linkage disequilibrium P value > 0.05), most likely due to recombination over time. Our results also suggest that both regions of class II genes may be included in the development of new genotyping methods based on next-generation DNA sequencing technologies.

Identification of cattle carrying alleles associated with resistance and susceptibility to the Bovine Leukemia Virus progression by real-time PCR

Previous studies have shown a significant association between polymorphisms of the BoLA DRB3 gene and Bovine Leukemia Virus (BLV) infection profile. The presence of allele ∗1501 has been associated with high proviral load in peripheral blood while allele ∗0902 has been associated with low proviral load. The purpose of this study was to develop allele-specific real-time PCRs to identify cattle carrying alleles associated with resistance (BoLA DRB3∗0902) or susceptibility (BoLA DRB3∗1501) to the BLV progression. Specific primers were designed and differential amplification was carried out by real-time PCR and monitored by SYBR® Green dye in DNA samples from peripheral blood. Conditions were also adjusted for traditional PCR amplification (end point amplification). These methods are rapid, simple and suitable for high throughput screening, and could aid in marker-assisted selection of BLV-resistant and susceptible cattle.

Identification of bovine leukocyte antigen class II haplotypes associated with variations in bovine leukemia virus proviral load in Japanese Black cattle

Bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leukosis, which is the most common neoplastic disease of cattle. Bovine leukocyte antigen (BoLA) is strongly involved in the subclinical progression of BLV infections. Recent studies show that the BoLA-DRB3 gene might play a direct role in controlling the number of BLV-infected peripheral B lymphocytes in vivo in Holstein cattle. However, the specific BoLA class II allele and DRB3-DQA1 haplotypes determining the BLV proviral load in Japanese Black cattle are yet to be identified. In this study, we focused on the association of BLV proviral load and polymorphism of BoLA class II in Japanese Black cattle. We genotyped 186 BLV-infected, clinically normal cattle for BoLA-DRB3 and BoLA-DQA1 using a polymerase chain reaction-sequence-based typing method. BoLA-DRB3*0902 and BoLA-DRB3*1101 were associated with a low proviral load (LPVL), and BoLA-DRB3*1601 was associated with a high proviral load (HPVL). Furthermore, BoLA-DQA1*0204 and BoLA-DQA1*10012 were related to LPVL and HPVL, respectively. Furthermore, we confirmed the correlation between the DRB3-DQA1 haplotype and BLV proviral load. Two haplotypes, namely 0902B or C (DRB3*0902-DQA1*0204) and 1101A (DRB3*1101-DQA1*10011), were associated with a low BLV proviral load, whereas one haplotype 1601B (DRB3*1601-DQA1*10012) was associated with a high BLV proviral load. We conclude that resistance is a dominant trait and susceptibility is a recessive trait. Additionally, resistant alleles were common between Japanese Black and Holstein cattle, and susceptible alleles differed. This is the first report to identify an association between the DRB3-DQA1 haplotype and variations in BLV proviral load.

Variation of the BoLA-DRB3 gene in dairy cattle and its effect on the viability parameters

Genotyping of the BoLA-DRB3 alleles was performed in dairy cattle of Krasnodar krai and Holstein stud bulls. Loss of heterozygosity, which decreased the reproductive parameters, was observed. It was proposed that stud bulls be selected on the basis of their genotyping at the BoLA-DRB3 gene to prevent further decay of cattle viability.

BoLA-DRB3 gene polymorphism and FMD resistance or susceptibility in Wanbei cattle

For the further characterization of foot-and-mouth disease virus (FMDV)-induced foot-and-mouth disease, we investigated the association between polymorphism of BoLA-DRB3 gene and FMD resistance/susceptibility of Wanbei cattle challenged with FMDV. One hundred cattle were challenged with FMDV and exon 2 of BoLA-DRB3 genes was amplified by hemi-nested polymerase chain reaction from asymptomatic animals and from animals with FMD. PCR products were characterized by the RFLP technique using restriction enzymes Hae III. The results revealed extensive polymorphisms, 6 RFLP patterns were identified. By analyzing alleles and genotypic frequencies between healthy and infection with FMD cattle, we found that allele Hae III A was associated with susceptibility to FMD in Wanbei cattle (P < 0.05), whereas Hae III C was associated with resistance to FMD (P < 0.01) and may have a strong protective effect against FMD. Hae IIICC and Hae III BC genotype were associated with resistance to FMD (P < 0.01). By contrast, Hae III AA genotype was associated with susceptibility to FMD (P < 0.01). Sequence analysis show that 89 amino acids were translated in exon 2 of BoLA-DRB3 and 13.70 % of nucleotide mutated, which resulted in 14.61 % of amino acid change. One PKC, one Tyr and one CAMP phosphorylation were increased; the hydrophobicity and secondary structure of proteins produced change after amino acid substitution. These results revealed that Wanbei cattle had the ability of resistance to disease by mutation which result changes of the protein structure to perform the regulation of the cell using different signaling pathways in the long process of choice evolution.

Identification of four novel alleles of the BoLA‐DRB3 upstream regulatory region in Chinese yellow cattle

The sequence of upstream regulatory region (URR) of BoLA-DRB3 gene was amplified with polymerase chain reaction followed by DNA sequencing from six animals of Chinese yellow cattle. A total of five alleles including four newly identified ones, named BoLA-DRB3*R-03-U2, BoLA-DRB3*R-06-U2, BoLA-DRB3*R-07-U and BoLA-DRB3*R-12-U for the BoLA-DRB3 URR were found. Result of sequence analysis showed that the regulatory elements W, X, Y, CCAAT and TATA-like boxes existed in such URRs and 16 polymorphic sites (11 transitions, 3 transversions, 1 deletion and 1 insertion) located in the spacers between the conserved consensus boxes and 1 insertion within X box, while no new polymorphic site within the consensus boxes.

Polymorphism of exon 2 of BoLA-DRB3 gene and its relationship with somatic cell score in Beijing Holstein cows

In the present study, the exon 2 and 3' end sequence of intron 1 of BoLA-DRB3 gene of 211 Beijing Holstein cows was amplified and a uniform fragment of 284 bp was obtained. The genetic polymorphism was investigated by PCR-RFLP using two restriction endonucleases HaeIII and BstYI. Seven genotypes were detected by digesting the PCR products with HaeIII. The frequency of AA, AB, AC, AD, BB, BC and BF genotypes was 0.4638, 0.0193, 0.0193, 0.3478, 0.0290, 0.0386 and 0.0822, respectively. Three genotypes were found by digesting the PCR products with BstYI. The frequency of AA, AB and BB genotypes was 0.0569, 0.2844 and 0.6587, respectively. The relationship between the polymorphisms in exon 2 of BoLA-DRB3 gene and somatic cell score (SCS) in Beijing Holstein cows was analyzed by least squares linear model. No significant difference was detected among least squares means of SCS for seven HaeIII-RFLP genotypes (P > 0.05). As for BstYI-RFLP analysis, least squares mean of SCS for AA was significantly lower than that for AB (P < 0.05) or BB (P < 0.05), and no significant difference was detected between AB and BB genotypes (P > 0.05). BstYI AA was the most favorable genotype and BstYI BB was the most unfavorable genotype for mastitis resistance. The information found in the present study is very important for improving mastitis resistance in dairy cattle by marker assisted selection.

Bovine MHC class II DR molecule plays a key role in bovine leukemia virus (BLV)-induced lymphoma

BLV is the etiological agent of enzootic bovine leukosis (EBL), which emerges as B-cell lymphomas. Previously, we found one of tumor-associated antigens (TAAs), which is a useful tool not only for diagnosing EBL but also for screening of BLV-infected cattle with the potential to develop tumor in the future, by using the monoclonal antibody (MAb) c143. We here performed biochemical characterization of c143 TAA such as two-dimensional electrophoresis, sequential immunoprecipitation and tryptic peptide mapping with c143 MAb as well as MAbs against bovine major histocompatibility complex (BoLA) class II antigens, and flow cytometry and immunofluorescence staining of cells transiently transfection with a combination of BoLA-DRA, -DRB3, -DQA and -DQB cDNAs. First, it was found that the c143 TAA is BoLA-DR consisting of alpha and beta chains. Furthermore, the altered expressions of c143 TAA were observed in infected animal with lymphoma at various stages as follows: 1) The number of cells positive for the c143 TAA and 2) glycosylation of the beta chain increased with the progression of EBL. 3) The beta chain was specifically phosphorylated at serine residue(s) during the lymphoma stage. Finally, we tested an association of microsatellite markers using 86 cattle with lymphoma and 320 asymptomatic cattle. We identified that seven genomic markers which located in chromosomes 1, 3, 9, 15 , 21, 23 and 24, and, in addition, BoLA-DRB3 gene in chromosomes 23, showed suggestive linkage to development of the BLV-induced EBL. Collectively, our results strongly indicate that BoLA-DR plays a key role in BLV-induced diseases progression.

Polymorphism of BoLA-DRB 3.2 gene in Iranian native cattle by using PCR-RFLP method

Enhancing disease resistance in animal production can be achieved by genetic improvement programs. The best characterized genetic control of disease resistance and immune response in animals is that associated with the MHC. Respond to specific antigens is closely related to MHC genes and their associations might therefore provide precious answers to main questions about the host pathogen interactions. In cattle there is an association between possession of certain BoLA-DRB3 locus (MHC class II) and resistance or susceptibility to infectious diseases. Polymorphism of exon 2 of the BoLA-DRB3 gene was investigated by the PCR-RFLP method in 80 Iranian native cattle. In this study, 39 different alleles were recognized. As a second goal, allelic frequencies were determined in a total sample. The high frequencies of these alleles (gbb, fbd and fba) were 8.75, 8.75 and 6.25% respectively. Additionally, BoLA-DRB3.2*11, DRB3*2701 and DRB3*0701 were associated with resistant to BLV, whereas allele DRB3.2*01 was associated with higher stability to FMD. Finally, DRB3*2701 and DRB3*1101 alleles were also associated with higher mastitis risk. Frequencies of alleles mediating resistance and susceptibility to leukemia and FMD were revealed. Key words: Iranian native cattle, MHC, BoLA-DRB3, PCR-RFLP.

Analyses of single nucleotide polymorphisms and haplotypes of BoLA-DRB3 gene in Holstein and Hanwoo

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→175/22), having one more cutting site by BstYI enzyme than d type allele and n' type (180→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.

Polymorphism of the BoLA-DRB3 gene in the Mongolian, Kalmyk, and Yakut cattle breeds

Polymorphism of the BoLA-DRB3 gene was studied with the use of the PCR-RFLP technique in three cattle breeds (Mongolian, Kalmyk, and Yakut) representing the Bos taurus turano-mongolicus group. 35 BoLA-DRB3.2 alleles were detected in the Mongolian breed and 34 alleles in the Kalmyk breed. The frequencies of alleles in both populations are distributed rather evenly: the frequencies of the most widely represented alleles (*18, *20, and *28) in the Mongolian cattle varied from 7.75 to 8.45%. The most frequent alleles in the Kalmyk cattle were *28(14.52%), *24(7.26%), and *12(6.45%). Only five alleles were identified in the Yakut cattle breed. The prevailing allele was *29 (77.3%); a relatively frequent allele was *1(13.1%), and the remaining three alleles constituted only 9.6%. Such a low level of diversity of BoLA-DRB3 gene alleles was not observed earlier in any other cattle breed. The Mongolian and Kalmyk breeds showed a wide diversity of BoLA-DRB3 genotypes (56 and 51 genotypes, respectively) and a high level of expected heterozygosity (He = 0.953 and 0.946, respectively). Both breeds had a deficiency of heterozygotes (Mongolian cattle: Ho = 0.775, D = -0.187; Kalmyk cattle: Ho = 0.708, D= -0.252). A low level ofgenotypic diversity for the BoLA-DRB3 locus (only seven genotypes; the frequency for the genotype *29/*29 is 71.4%) and a very low level of observed heterozygosity (Ho = 0.12) were revealed in the Yakut breed. BoLA-DRB3.2 alleles associated with resistance to persistent lymphocytosis caused by the bovine leukemia virus (total frequencies 15.49 and 24.19%) and to various forms of mastitis (total frequencies 12.68 and 20.96%, respectively) were identified in the Mongolian and Kalmyk animals. In the Yakut breed, alleles associated with resistance to diseases are represented only by the BoLA-DRB3.2 allele *7 (1.2%). Thus, the Mongolian and Kalmyk cattle breeds are characterized by a wide diversity of alleles and genotypes for the BoLA-DRB3 gene. In contrast, the population of Yakut cattle from the Verkhoyanskii raion of the Republic of Sakha has a poor diversity of alleles and genotypes for the BoLA-DRB3 gene and a very low level of heterozygosity, suggesting an unfavorable state of the population that is probably caused by inbreeding depression due to a long-term isolation and a small number of animals.

Restriction fragment length polymorphism in the exon 2 of the BoLA-DRB3 gene in Chinese Holstein of the south China

The genetic diversity of the exon2 of BoLA-DRB3 (BoLA-DRB3.2) in Chinese Holstein cattle of the south China was investigated by hemi-nested polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Six, four and eleven RFLP patterns were found after digestion with the restriction enzymes Hae III, Bst YI and Rsa I, respectively. The DNA sequence showed and twenty- five DRB3.2 alleles. GLM model analysis indicated that lactation and calving season have positive correlation with SCC (Somatic Cell Count) (p<0.01), BoLA-DRB3.2*3, *8, *18 and *26 were associated with lower SCC (p<0.01). The present findings concluded that mastitis is a severe hinder of milk production and technology. Therefore, future re-searches should focus on associations of mastitis with BoLA haplotypes rather than single BoLA genes

Association of BoLA DRB3 and DQA1 alleles with susceptibly to Neospora caninum and reproductive outcome in Quebec Holstein cattle

The BoLA DRB3 and DQA1 genes are part of the major histocompatibility complex (MHC) class II in cattle. These genes are highly polymorphic and have been associated with resistance to several diseases, such as mastitis, Bovine Leukemia Virus (BLV) and dermatophilis. Sequenced based typing of these genes has been carried out extensively from blood samples; however it is often impractical or expensive to obtain such samples. Repositories of well-characterized serum from cattle are readily available in many veterinary research facilities. This paper reports a retrospective analysis of BoLA class II genotypes of cattle obtained from stored serum samples from Holstein cattle from Québec dairy farms, which were obtained as part of a previous study on bovine neosporosis. It was possible to genotype 56 cattle with known infection status for Neospora caninum. We identified 14 different DRB3 and 10 different DQA1 alleles in this population. The allele frequency distribution was consistent with previously studied cattle populations, and alleles known to be associated with BLV and mastitis were present. No association was found between allele frequency distribution of DRB3 or DQA genes and infection with N. caninum. However, an association of allele DRB3*1001 and allele DRB3*2703 with resistance and susceptibility to pregnancy loss, irrespective of infection status, was identified.

Distribution of BoLA-DRB3 allelic frequencies and identification of a new allele in the Iranian cattle breed Sistani (Bos indicus)

The distribution of the frequencies of BoLA-DRB3 gene alleles in the Iranian cattle breed Sistani was studied by the PCR-RFLP ("hemi-nested") assay using restriction endonucleases RsaI, HaeIII and BstYI. In the examined cattle breed (65 animals) 32 alleles have been identified one of which being described for the first time (6.15% frequency). The nucleotide sequence of the polymorphic region of exon 2 of this allele has been determined and submitted in the GeneBank database under accession number DQ486519. The submitted sequence has maximum homology (92%) with the previously described sequence DRB3-mRNA from Bos indicus (AccN X79346) and differs from it by 24 nucleotide substitutions which result in 16 amino acid substitutions. The peptide (on the basis of the reconstructed amino acid sequence) has 89% identity to the sequence encoded by the BIDRBF 188 locus (Bos indicus). The results obtained permit the sequence described by us to be considered as a new allele of the BoLA-DRB3 gene (DRB3.2**X). The total frequency of the main six alleles (DRB3.2*X, *10, *11, *20, *34 and *X) occurring with a frequency of over 5% is about 60% in Iranian Sistani cattle. Fifteen alleles have <1% frequency. The highest frequency was observed for DRB3.2*8 allele (21.54%) like in other previously described breeds of Bos indicus (up to 23.07%). The Iranian breed Sistani has a high level of similarity by the spectrum of BoLA-DRB3 alleles and their frequencies to other Bos indicus breeds and significantly differs by these criteria from the Bos taurus breeds. The Iranian Sistani herd under study includes alleles associated with to resistance to leukemia (DRB3.2*ll and *23) and to different forms of mastitis (DRB3.2*2, *7, *11, *23 and *24) although their frequencies are low (from 0.77 to 5.37%). On the whole, a high level of diversity of BoLA-DRB3 gene alleles and the availability of alleles associated with resistance to different diseases makes this breed of interest for breeding practice.

Evidence for cattle major histocompatibility complex (BoLA) class II DQA1 gene heterozygote advantage against clinical mastitis caused by Streptococci and Escherichia species

Mastitis is an inflammatory response of the mammary gland to irritation, injury, or infectious agents and is a major problem in the dairy industry. We genotyped bovine major histocompatibility complex (BoLA)-DRB3 and BoLA-DQA1 genes in 120 Holstein cattle with clinical mastitis and 85 randomly selected Holstein cattle in Japan by polymerase chain reaction-sequence-based typing. The mastitis cattle were divided into four groups according to the bacterial species that caused the mastitis (Staphylococcus aureus, Streptococci, Escherichia, and coagulase-negative staphylococci). The BoLA-DRB3 and BoLA-DQA1 heterozygosity of each group was compared with that of the control cattle, while the expected heterozygosities based on Hardy-Weinberg proportions and the observed heterozygosities for each locus were compared for each group. The Escherichia-induced and Streptococci-induced mastitis groups showed significant differences between their expected and observed heterozygosities with regard to their BoLA-DQA1 genes. No differences were observed for any group with regard to the BoLA-DRB3 genes. We then found that two BoLA-DQA1 alleles promoted susceptibility to Streptococci-induced mastitis, namely BoLA-DQA1*0101 and BoLA-DQA1*10012 and that the homozygous BoLA-DQA1*0101/0101 and BoLA-DQA1*10011/10011 genotypes promoted susceptibility to mastitis caused by Streptococci and Escherichia, respectively. This is the first report showing that heterozygosity of the BoLA-DQA1 gene is associated with resistance to mastitis progression.

Association of the &amp;lt;i&amp;gt;BoLA-DRB3&amp;lt;/i&amp;gt; alleles with estimated breeding value for somatic cell count in Polish dairy cattle

Abstract. Introducing resistance to mastitis into breeding programmes for dairy cattle seems to be one of the possible methods for limiting the increasing number of mastitis cases. Looking for candidate genes for genetic markers is one of the strategies. The BoLA–DRB3 alleles have considerable promise as potential mastitis marker. The objective of the study was to evaluate relationships between two BoLA–DRB3 alleles (BoLA–DRB3.2*16 and BoLA-DRB3.2*23) and estimated breeding value (EBV) for somatic cell count (SCC) in Polish dairy cattle. A total of 525 Polish Holstein cows in two experimental farms were evaluated for test-day SCC. The polymorphism of BoLA–DRB3 gene was identified using the MPT-PCR method. The variance components for test-day SCC were estimated by the REML method using animal model. The breeding value of SCC was calculated according to the BLUP method. Linear contrast between model-adjusted breeding values of SCC for all animals was used to test for differences between genotypes. In Polish Holstein cows population BoLA allele DRB3.2*23 was associated with increase of EBV for SCC in milk. It was observed increase of EBV for SCC from cows carrying this allele comparing to cows carrying BoLA–DRB3.2*16 (p ≤0.07) or other alleles (p ≤0.01).

A rapid and robust sequence‐based genotyping method for BoLA‐DRB3 alleles in large numbers of heterozygous cattle

The BoLA-DRB3 gene is a highly polymorphic major histocompatibility complex class II gene of cattle with over one hundred alleles reported. Most of the polymorphisms are located in exon 2, which encodes the peptide-binding cleft, and these sequence differences play a role in variability of immune responsiveness and disease resistance. However, the high degree of polymorphism in exon 2 leads to difficulty in accurately genotyping cattle, especially heterozygous animals. In this study, we have improved and simplified an earlier sequence-based typing method to easily and reliably genotype cattle for BoLA-DRB3. In contrast to the earlier method, which used a nested primer set to amplify exon 2 followed by sequencing with internal primers, the new method uses only internal primers for both amplification and sequencing, which results in high-quality sequence across the entire exon. The haplofinder software, which assigns alleles from the heterozygous sequence, now has a pre-processing step that uses a consensus of all known alleles and checks for errors in base calling, thus improving the ability to process large numbers of samples. In addition, advances in sequencing technology have reduced the requirement for manual editing and improved the clarity of heterozygous base calls, resulting in longer and clearer sequence reads. Taken together, this has resulted in a rapid and robust method for genotyping large numbers of heterozygous samples for BoLA-DRB3 polymorphisms. Over 400 Holstein-Charolais cattle have now been genotyped for BoLA-DRB3 using this approach.

The Diversity of BoLA-DRB3 Gene in Iranian Native Cattle

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.

Comparison of bovine lymphocyte antigen DRB3.2 allele frequencies between two subpopulations of Iranian holstein cattle

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 favor the binding of different peptides, DRB3 has been extensively evaluated as a candidate marker for associations with various bovine diseases and immunological traits. Therefore, in this study, the genetic diversity of the bovine class II DRB3 locus in the two Iranian subpopulations of Holstein cattle (Moghan farm n = 250 and Razavi farm n = 175) was investigated by polymerase chain reactionrestriction fragment length polymorphism method (PCR-RFLP). Bovine DNA was isolated from whole blood. A hemi-nested PCR followed by digestion with restriction endonucleases RsaI, HaeIII and BstYI was conducted on the DNA. The results indicated that exon 2 of the BoLA-DRB3 gene is highly polymorphic in the two populations, and the frequency of BoLA-DRB3 depends on breed. On the other hand the presence of BoLA-DRB3*8, *24, and *16 alleles with high frequency in BoLA-DRB3.2 system, can be used as appellative index for nominate Holstein.