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Genetic Relationship Among Indigenous Sheep Population of Bangladesh

GK Deb
Bangladesh Livestock Research Institute, Savar, Dhaka

MP Choudhury
Bangladesh Livestock Research Institute, Savar, Dhaka

MA Kabir
Bangladesh Livestock Research Institute, Savar, Dhaka

MYA Khan
Bangladesh Livestock Research Institute, Savar, Dhaka

M Ershaduzzaman
Bangladesh Livestock Research Institute, Savar, Dhaka

TN Nahar
Bangladesh Livestock Research Institute, Savar, Dhaka

SMJ Hossain
Bangladesh Livestock Research Institute, Savar, Dhaka

MS Alam
Bangladesh Livestock Research Institute, Savar, Dhaka

MA Alim
National Institute of Biotechnology, Savar, Dhaka

Abstract/ Executive Summary:

The study was conducted to investigate the genetic relationships among indigenous sheep population of Bangladesh (Barind, Jamuna river basin, Coastal and Garole sheep) using microsatellite markers. A total of 96 blood samples were collected from adult sheep of Barind (24), Jamuna River Basin (24), Coastal (24), Garole(10) and available Chotanagpuri (10) sheep. Chotanagpuri sheep was used as an out group population. DNA was extracted from blood samples using QIAGEN DNA Mini extraction kit and was quantified using a nanodrop. FAO recommended 13 labeled microsatellite markers were used for polymerase chain reaction (PCR). PCR product was confirmed by 2% agarose gel electrophoresis and visualized by staining with ethidium bromide.The exact allele sizes in each primer were determined by GeneMaker V1.85 demo. Microsatellite tool kit and Dispan software package were used for calculation of allele frequency, number of alleles per locus, observed and expected heterozygosity and genetic distances (D_A). The Dispan software was used to calculate inter-individual genetic distances. These distance values were used to construct an UPGMA tree. Results showed that average number of polymorphic alleles per locus varied from 4 in HUJ616 to 12 in MAF70. Observed heterozygosity was also varied from 0.54±0.04 in Coastal to 0.63±0.03 in Barind sheep population. Genetic distance between Jamuna river basin and Barind was lowest (0.01) and between Garole and Costal was highest (0.17). Garole and Chotonagpuri sheep has higher genetic distance from other three sheep populations. Phylogenetic dendogram showed that sheep of Jamuna river basin and barind were belonged to same genetic group. Whereas, coastal, garole and Nagpur sheep were shown higher genetic distances from Jamuna river basin and coastal sheep. Considering findings of this study it may be concluded that the Barind and Jamuna river basin sheep belongs to a similar genetic group while, Garole and Coastal sheep are belonging to two distinct genetic groups.

Keywords: Dendogram; Genetic distances; Heterozygosity; sheep

Location: Genetics Laboratory of Bangladesh Livestock Research Institute

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Sheep

Objectives:

To evaluate genetic relationship among different sheep population of Bangladesh using microsatellite markers.

Methodology:

Based on geographic distribution, management system and breeding history, a total of 96 blood samples were collected from five sheep population of Bangladesh. The populations were Barind (Naogaon, Rajshahi and Chapainowabganj district), Jamuna River Basin (Tangail, Sirajgonj and Mymensingh district), Coastal (Noakhali district), Chotonagpuri (Meherpur district) and Garole (Khulna district) breeds. Samples were collected only from adult sheep of subsistent farmers and BLRI flock. Samples were collected avoiding related animals. Blood was collected in venoject tube, treated with anticoagulant and carried to Molecular Genetics Laboratory of BLRI and preserved at -20^oC until DNA extraction. DNA was extracted using a commercial kit (QIAGEN DNA Mini Kit) following manufacturer’s instructions.DNA samples were quantified using Nanodrop 2000 spectrophotometer. Thirteen FAO recommended microsatellite primer pairs were used (FAO, 2011). Most of primers used were independent and belonged to different chromosomes. All microsatellite markers were forwardly labeled with a capillary based dye: FAM (blue), PET (Red), VIC (Green) and NED (yellow) for the purpose of genotyping. PCR amplifications were performed with a programmable thermal cycler (Bio-Rad, DNA Engine, T100) in a final reaction volume of 25 µL using PCR reagents from Invitrogen. Two types of PCR reaction mixture were used. The first reaction system consisted of 2 µL of 50 ng of genomic DNA, 0.5 µL of each primer, 2.5µL 10X PCR buffer, 2µL dNTPs, and 0.937 U Taq DNA polymerase [TaKaRa Biotechnology (Dalian) China] and the second one consisted of 2 µl containing 50 ng genomic DNA, 1 µl of each primer, 12.5µl premix and 8.5 µl ddH₂O. The amplification conditions were as follows: 5 min at 94ºC for initial denaturing followed by 35 cycles. at 94ºC for 30 s; annealing for 30 s; extension at 72ºC for 30 s and a final extension at 72ºC for 7 min for all primers. Amplification was confirmed by gel electrophoresis of the PCR products on 2% agarose gel followed by visualization under UV. PCR products with different florescent dye were subjected to capillary electrophoresis using Gene Scan -500 LIZ Size Standard with DNA Analyzer 3730 of Applied Biosystemin Life Biotech Ltd. China. The exact allele sizes in each primer were determined by Gene Maker V1.85 demo Genotypes were assigned for each animal based on allele size data. Allele frequency, the mean number of alleles per locus, observed heterozygosity, and heterozygosity expected from Hardy-Weinberg assumptions for each locus, and Nei’s D_A genetic distances (Nei, 1983) were computed using the Microsatellite tool kit and Dispan software package. The Dispan Computer program was used to calculate inter-individual genetic distances, based on the proportion of shared alleles. These distance values were used to construct an unweighted pair group-method with arithmetic mean (UPGMA) phylogenetic tree using Dispan software (Saitou and Nei, 1987).

Source of Information: Bang. J. Anim. Sci. 2019. 48 (1): 17-22

Article Link (Online Journal): DOI: https://doi.org/10.3329/bjas.v48i1.44553

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Considering findings of the present study on average allele numbers, heterozygosity and standard genetic distances and dendrogram, it may be concluded that the Barind and Jamuna river basin sheep are belonging to a similar genetic group while, Garole and Coastal sheep are two distinct genetic groups.

Knowledge Management: Journal