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Rapd-based Genotyping of Walking Catfish, Clarias Batrachus in a Population of Bangladesh

Md. Faruque Miah
Department of Genetic Engineering and Biotechnology,Shahjalal University of Science and Technology, Sylhet, Bangladesh

Toufiqur Rahman
Department of Genetic Engineering and Biotechnology, Shahjalal University of Science and Technology, Sylhet, Bangladesh

Shahena Aktar Shipa
Department of Genetic Engineering and Biotechnology, Shahjalal University of Science and Technology, Sylhet, Bangladesh

Anindita Chakrabarty
Department of Genetic Engineering and Biotechnology, Shahjalal University of Science and Technology, Sylhet, Bangladesh

Abstract/ Executive Summary:

Genetic diversity of any fish is most important for their existing, breeding, production and management. In this study, intra-species genetic variation of a wild population of walking catfish, Clarias batrachus in Sylhet, Bangladesh was investigated. Four RAPD markers such as OPB-19, OPB-12, OPB-05 and OPF-14 were used to assess genetic variability among 8 individuals of C. batrachusgenotypes. Less number of DNA bands was revealed by all the primers in experimental populations where a total of 132 bands were detected and all the markers were shown different levels of polymorphism. Intra-specific polymorphisms were recorded 60%, 55.56%, 40.91% and 30.77% in primer OPB-12, OPB-19, OPF-14 and OPB-05 respectively where average polymorphism was recorded 46.81%. Average gene diversity was found 0.19823. The genetic distance of experimental samples ranged from 0.6 to 1 and in an average 0.88, indicating higher genetic distances of this population. A genetic relationship among the individuals of the experimental population was constructed where 3 clades and 2 sub-clusters were recorded considering different linkage distances. Considering different genetic parameters genetic variability was found higher among this experimental population.

Keywords: Genetic diversity, Walking catfish, RAPD, Clarias batrachus

Location: Sylhet, Bangladesh

Start Date:

End Date:

Program Area: Conservation and Biodiversity

Commodity: Cat fish

Objectives:

In this study, a cost-effective and simple RAPD technique was analyzed to assess the genetic diversity in a wild population of C. batrachus in Bangladesh

Methodology:

Sample collection The experimental samples of fish samples were collected by traditional fish catcher from haor habitat (a type of freshwater) of Sylhet, Bangladesh, which were then brought to the Fisheries and Marine Biotechnology Research Unit in the Department of Genetic Engineering and Biotechnology (GEB) at Shahajalal University of Science and Technology (SUST), Sylhet, Bangladesh. The collected fish samples were identified through morphometric characteristics of Shafi and Quddus (1982) and Rahman (1989) and kept in aquariums by maintaining the proper environment until tissue isolation. DNA Extraction The fishes were dissected to take out the liver and kidney by using scissors, forceps and needles. The isolated tissues of the fishes were taken separately into the petridish and washed with distilled water. Then the organs were kept in falcon tubes using 70% ethanol and preserved them at -20ºC until DNA extraction. DNA extraction was done from tissues using a commercially available kit, Gene JET and the quality of extracted DNA was checked by gel electrophoresis using 0.8% agarose. After that, the DNA samples were stored at-20ºC. PCR amplification Genotyping of Asian catfish, C. batrachusin a population of Bangladesh was studied using four decamer primers and the selected primers were OPB-19, OPB-12, OPB-05 and OPF-14. PCR reaction was carried out using GoTaq® G2 Hot Start Green Master Mix and 15 μl PCR mixtures were used for each DNA sample. PCR reaction was conducted with pre-heating at 94⁰C for 3 minutes followed by 35 cycles of denaturation at 94ºC for 1 minute; annealing temperature for selected four primers OPB-19, OPB-12, OPB-05 and OPF-14 were 29ºC, 27ºC, 29ºC and 30ºC respectively for 1 minute and 2 minutes elongation or extension at 72⁰C. A final step of 7 min for 72ºC was added to allow complete extension of the amplified fragments. The amplified PCR products from each sample were checked for the banding pattern of DNA by electrophoresis on 1.2% agarose and gel DNA was contained using ethidium bromide. A ladder (1kb Gene Rullar, USA) ranging 250bp-10000bp was used for comparison of DNA quality. The agarose gel of the amplified PCR products was placed on the gel documentation to take the photograph (Panasonic” DMC-FS20, 10 Megapixels). Assessment of genetic variability RAPD data of this experiment was interpreted by using different software and equations whereas Alpha Ease FC 4.0 was used for the identification of bands at molecular level. Polymorphism information content (PIC) was measured using a formula where the PIC is 2fi(1-fi) where a sfi is the frequency of the amplified allele (band present). The average number of alleles per locus was calculated by a formula.

Source of Information: Journal of Entomology and Zoology Studies 2020; 8(1): 885-888 E-ISSN: 2320-7078P-ISSN: 2349-6800

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

In this experiment, RAPD based genetic variability of wild type Clarias batrachus in Sylhet, Bangladesh was analysed. Based on this study considering different genetic parameters higher genetic variability was found in this experimental population. However, it is very much needed for observing genetic status of this fish with more individuals from different habitats with different markers whereas the genome of this fish is already sequenced. However, this finding would be used for the future investigation of this species in Bangladesh.

Knowledge Management: Journal