Siraj, Siti Shapor

Tan, Soon Guan

Yusoff, Khatijah

Universiti Putra Malaysia Institutional Repository

UPM Research Report 1997-2000. Vol II.Section 2-Extended AbstractsApplication of Microsatellite Markers for Stock Identification and Improvement ofthe Malaysian River Catfish Mystus nemurusTan. S. G.. SlrsJ. S.S ••Yusoff. K.Facultyof Science and Environmental Studies,Universiti Putra Malaysia43400 UPM, Serdang, SelangorMalaysiaTelephone Number of Corresponding Author: 03-89466640E-mailolCorrespondingAuthor:sgtan@ftas.upm.edu.myKey words: river catfish, DNA, genetic markers, characterizationIntroductionThe river catfish Mystus nemurus is widely distributed in both mainland as well as archipelago Southeast Asia. Due to its goodflesh quality it is a popular fresh water food fish throughout the region and is therefore of interest to aquaculturists. Earlier wetyped five populations of river catfish from Peninsular Malaysia (Kedah, Perak, Selangor, Johor, Terengganu) for 16 locicoding for nine enzymes and sarcoplasmic protein. These populations were also typed for DNA markers by using the RandomAmplified Polymorphic DNA (RAPD) and the Amplified Fragment Length Polymorphism (AFLP) techniques. Another classof DNA markers called microsatellites are very powerful genetic tools. They are codominant and have become increasinglypopular as genetic markers for use in a variety of applications. Our aim is to isolate and characterize microsatellite markers forthis commercially viable species.Materials and MethodsRiver catfish samples from Perak, Kedah, Johor, Selangor and Sarawak were collected and brought back to our laboratory. Formicrosatellite isolation only one DNA sample was required. Microsatellite loci were isolated using two techniques. The firsttechnique involved using RAPD primers and cloning of the PCR products foIlowed by screening of the clones formicrosatellites. This procedure is called RAHMs. The second procedure called Random Amplified Microsatellite (RAM) is aS' anchored PCR technique used to isolate microsatellites. Both these procedures are shortcuts to the conventional methodwhich have proven to be efficient and economical. The microsatellite loci that we developed are being used in a follow upproject on gene mapping of this species. Primers were designed for each microsatellite locus and tested for polymorphism.Direct Amplification of Length Polymorph isms (DALP) was also used to obtain codominant DNA markers for this species inthe absence of DNA sequence data. Six primers have been assessed for the level of polymorph isms among the fivepopulations.Results and DiscussionMicrosatellites allow accurate genetic assessment of population differentiation. They are the most frequently used type ofmarkers in linkage mapping. They are codominant markers and are known to be associated with quantitative trait loci. Thiswould prove beneficial for the aquaculture industry as the breeders can then make informed decisions on their brood stocks.The use of RAHMs and RAMs together resulted in the isolation of over 100 microsatellite loci in M nemurus. A total of 26simple sequence repeats were identified using RAHMs. Five of these have been characterized. The average number of allelesper locus was 3.2. Primers have also been designed for another 56 loci and are being tested for polymorph isms. Till now wehave found 30 polymorphic markers. These markers can be tested for possible associations with quantitative traits of economicimportance.ConclusionsCodominant microsatellites are powerful genetic markers which are useful for the precise characterization of the species thatcan help in choosing the right stocks for breeding programmes. The microsatellite loci which were developed during the courseof this project are now being mapped in a follow up project on gene mapping entitled Population Characterization andLinkage mapping in Ikan Baung, Mystus nemurus, using Microsatellite Markers which is part of a Malaysia-Thailand BilateralBiotechnology Project on Genome Mapping of Freshwater Catfish.Benefits from the studyKnowledge as reported in scientific publications which includes journal, conference and seminar papers as well as theses ofundergraduate and postgraduate students. This project started in 1996 and was extended with additional funding in 1999.Patent{s), if applicable:NiI-Stage of Commercialization, if applicable:Nil-258Agricultural SciencesProject Publications in Refereed JournalsJ. Chong LK, Tan SG, YusofT K, Siraj SS. 2000 Identification and characterization of Malaysian river catfish, Mystus nemurus (C & V):RAPD and AFLP analysis. Biochemical Genetics 38: 63-752. Chong LK, Tan SG, Siraj SS, Christianus A and YusolT K. 1999. Mendelian inheritance of random amplified polymorphic DNA (RAPD)markers in the river catfish. Malaysian Applied Biology 28: 79-84.3. Usmani, 5., Tan, S. G., Siraj, S. S. &YusofT, K, (2001) Isolation and characterization of micro satellites in the Southeast Asian river catfishMystus nemurus. Molecular Ecology Notes I: 264-266.4. Usmani, S., Tan. S.G., Siraj, S.S., Yussof, K. (2003). Population structure of the Southeast Asian river catfish Mystus nemurus as revealedby twenty microsatellite markers. Animal Genetics (In press).Project Publications in Conference ProceedingsJ. Bhassu, S., YusofT, K., Panandam, 1.M., Embong, W.K., Sivarajasingam, S., Oyyan, S., Tan, S.G. (1999). Microsatellite analysis ofOreochromis spp. In: Proceedings Malaysian Science and Technology Congress '99. Confederation of Scientific and TechnologicalAssociations in Malaysia (COST AM). Kuala Lumpur. p 62-68.2. Usmani, S., Tan, S.G. (1999). Capture of microsatellites using direct amplification of length polymorphisms. In: Proceedings MalaysianScience and Technology Congress '99. Confederation of Scientific and Technological Associations in Malaysian (COSTAM). KualaLumpur. p, 439-444.3. Usmani, S., Chua, B.E., Tan, S.G., Siraj, S.S., YusofT, K.M. (2000). A study of three populations of the river catfish Mystus nemurus usingrandom hybridising microsatellites. In: Saad et al. (Eds). Genetic Manipulations: Challenges and Advances. Genetics Society of Malaysia.Bangi. p. 88-97.4. Usmani, S., Tan, S.G., Siraj, S.S., YusolT, K.M. (2001). Isolation of microsatellites in Mystus nemurus using 5' anchored PCR. In:Proceedings of The 6th Symposium of the Malaysian Society of Applied Biology, 12-13 February 2001 at Monash University Malaysia.Published on CD. Malaysian Society of Applied Biology, Bangi.5. Hoh, B.P., Tan, S.G., Siraj, S.S., YusofT, K. (2003). Identification of micro satellite loci in Mystus nemurus using RAPD markers. In : M.K.Thong (ed.) From Peas to Chips: The Globalisation of Genetics. The Genetics Society of Malaysian. Bangi. p. 45-46.Graduate ResearchName of Research Topic Field of Expertise Degree Awarded Graduation YearGraduateChong Lee Kim Development of Genetics MS 1999PCR-based DNAmarkers to identifyand characterizeMalaysian rivercatfish, Mystusnemurus (C & V):RAPD and AFLP.Sahar Usmani Isolation, Genetics PhD 2002characterization andapplication ofmicrosatellitemarkers in theSoutheast Asianriver catfish (Baung)Mystus nemurus (C&V).Subha Bhassu Applications of Genetics PhD 2002DNA microsatellitemarkers In tilapiaculture.259UPM Research Report 1997-2000. Vol II. Section 2-Extended AbstractsIRPA Project numberOl-02-04-0074UPM Research Cluster:AFFProject Leader Tan Soon Guan260

Application of microsatellite markers for stock identification and improvement of the Malaysian river catfish Mystus nemurus

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Application of microsatellite markers for stock identification and improvement of the Malaysian river catfish Mystus nemurus

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