تجزیه‌ و تحلیل ژنوتیپی ناحیه‌ی کمپلکس اصلی سازگاری بافتی در مرغ‌های بومی ایران

پژوهشهای علوم دامی ایران

مقاله 8، دوره 11، شماره 3 - شماره پیاپی 39، مهر 1398، صفحه 365-375   

نوع مقاله: علمی پژوهشی- ژنتیک و اصلاح دام و طیور

شناسه دیجیتال (DOI): 10.22067/ijasr.v11i3.71958

نویسندگان

جعفر پیش جنگ آقاجری^{* 1}؛ قدرت اله رحیمی میانجی²؛ سید حسن حافظیان³؛ محسن قلی زاده⁴؛ قربان الیاسی⁵

¹دانشگاه آزاد اسلامی، واحد مراغه

²گروه علوم دامی، دانشـکده ی علـوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران.

³گروه علوم دامی، دانشکده‌ی علوم دامی و شیلات، دانشگاه علوم کشاورزی و منابع طبیعی ساری، ساری، ایران.

⁴دانشگاه علوم کشاورزی و منابع طبیعی ساری

⁵مرکز تحقیقات کشاورزی و منابع طبیعی استان آذربایجان شرقی

چکیده

ناحیه­ی MHC مرغ‌ها در پاسخ‌های ایمنی، مقاومت به بیماری‌ها و فرایندهای تکاملی، دارای اهمیت است. در این تحقیق 200 نمونه­ی خونی از مرغ­های بومی نژاد­های عمومی، آذربایجان غربی، مرندی و مازندرانی اخذ شد و DNA­ی ژنومی به روش بهینه شده­ی نمکی استخراج و چند­شکلی­های­ آللی در جایگاه‌های ژنی B-L، B-F و B-G با استفاده از تکنیک PCR-RFLP بررسی شد. برای شناسایی جهش در جایگاه‌های ژنی یاد شده از آنزیم­ Msp I استفاده شد. در جایگاه ژنی 374 جفت بازی B-L، فقط ژنوتیپBB  اما در جایگاه ژنی 1048 جفت بازی B-F، دو ژنوتیپ CG و GG شناسایی شد. در این جایگاه آلل C شامل باندهای 515، 410، 75 و 47 جفت بازی و آلل G نیز دارای باندهای 410، 302، 213، 75 و 47 جفت بازی بودند. در جایگاه ژنی 401 جفت بازی B-G، سه ژنوتیپ MM، MN و NN و دو آلل M شامل یک باند 401 جفت بازی و آلل N دارای باندهای 350 و 51 جفت بازی مورد شناسایی قرار گرفت. در کل جمعیت­ها شاخص اطلاعات شانون در دو جایگاه ­ژنی B-F و B-G به ترتیب 37/0 و 59/0 و شاخص تثبیت به ترتیب 13/0- و 17/0- محاسبه شدند. بیشترین مقدار شاخص هتروزیگوسیتی مشاهده‌ شده برای جایگاه­های ژنی یاد شده به ترتیب 25/0 و 48/0 برآورد شد. با توجه به وجود چندشکلی در دو جایگاه ژنی B-F و B-G، می‌توان با استفاده از پاسخ ایمنی ژنوتیپ‌های مشاهده ‌شده، از این جایگاه‌ها به ‌عنوان نشانگر برای اصلاح نژاد ژنتیکی جهت افزایش مقاومت مرغ‌های بومی به بیماری­ها‌ استفاده کرد.

کلیدواژه‌ها

چندشکلی؛ مجموعه سازگاری بافتی (MHC)؛ مرغ بومی؛ PCR-RFLP

مراجع

Introduction:

Chicken major histocompatibility complex (MHC) region are important in immune responses, resistance to diseases, and relationships with evolution processes. The chicken major histocompatibility complex is composed of two gene regions: the B and Y (Rfp-Y) loci, both located on micro chromosome 16. The B locus includes three gene classes, I (B-F), II (B-L) and IV (B-G). The chicken major histocompatibility complex, consists of several clusters of highly polymorphic genes, some of which are associated with disease resistance. The class I and class II antigens resemble their mammalian counterparts in the encoded protein structure. The class IV region encodes the B blood group antigens, which are readily identified by serological blood-typing. The class III region appears to be divided in chickens, with some elements that are MHC-linked and others that map elsewhere. In addition the Rfp-Y system, which bears a strong similarity to the MHC, maps to the opposite side of the nucleolar organizer region on the same micro chromosome as the MHC. Each class of MHC genes is a potential candidate for a role in disease resistance. The MHC genes show associations with response to diseases as diverse as virally induced neoplasia, bacterial, parasitic and auto-immune diseases.

 

Materials and Methods:

In this study, allelic polymorphism in B-L, B-F and B-G loci involved in the immune system in four Iranian indigenous chickens were examined using PCR-RFLP technique. Two hundred birds including common, West Azerbaijan, Marandi, Mazandarani indigenous chicken breeds were selected. As much as 1-2 ml of blood was taken from each of the chicken. Blood samples were transferred to the anticoagulant (Ethylene diamine tetra acetic acid) tubes in the vicinity of the ice to the genetic and biotechnological laboratory of Islamic Azad University, Maragheh branch and until the onset of genomic DNA extraction and subsequent experiments were kept at -20°C. In the extraction of the genomic DNA of blood samples, the salting out method and for amplify of each locus, a pair of specific primers was used. For detection of mutation in the loci the Msp I enzyme was used. For genetic analysis of data derived from digestive enzymes in indigenous chickens, POPGENE software version 1.32 was used. This software is used to estimate the allele and genotypic frequencies, observed and expected heterozygosity, mean heterozygosity, Hardy-Weinberg equilibrium, Fixation index, Shannon information index, and other genetic parameters.

 

Results and Discussion:

According to this study results, in the 374-bp locus of B-L, after enzymatic digestion, only BB genotype and monomorphic was detected. In the 1048 bp locus of B-F, two genotypes CG and GG were identified and the C allele included 515, 410, 75 and 47 bp bands, and the G allele also included bands of 410, 302, 213, 75 and 47 bp and the χ2 calculated in this locus was not significant for all populations (P < 0.05), and all populations were in Hardy-Weinberg equilibrium. Three Genotype MM, MN and NN genotypes were identified for the locus of B-G (401 bp), M allele included a 401 bp band and N allele included bands of 350 and 51 bp. The χ2 calculated in this locus was not significant for the indigenous chicken population of Mazandarani (P < 0.05) and this population was in Hardy-Weinberg equilibrium. The Shannon information index was calculated to be 0.37 and 0.59 in markers loci of B-F and B-G, respectively, and the fixation index values were -0.13 and -0.17, respectively. The highest observed heterozygosity index for B-L and B-G loci was 0.24 and 0.57, respectively. Estimation of the negative fixation index values in the studied chickens populations could be due to the high selection rate in the populations. The fixation index values is always variable in range -1 to 1, and its negativity indicates a decrease in heterozygosity and increase in homozygosity or increased inbreeding, as well as a deviation from the Hardy-Weinberg equilibrium in the populations. Shannon's information index is an estimate of genetic diversity in populations. In all of the populations studied, the B-G locus has a relatively high genetic diversity.

 

Conclusion:

Regarding the polymorphism in the two gene sites (B-F and B-G) studied and the heterozygosity reduction in the populations studied, can be prevented from occurrence of non-random crosses in populations and prevented the reduction of heterozygosity and thus reduced genetic diversity. Also, by studying the immune responses associated with these two gene sites, from these genes can be used as marker for genetic breeding in indigenous chickens for increase of resistance to diseases.