PARENTAGE VERIFICATION IN FIELD PROGENY TESTING PROGRAMME OF MEHSANA BUFFALO BY MICROSATELLITE ANALYSIS
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Date
2008
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AAU, Anand
Abstract
The knowledge of correct parentage is a prerequisite in breeding programmes. The identification of proven sires has been of utmost importance in animal improvement programmes. Failure to record correct parentage can cause bias in sire evaluation, by introducing errors in estimates of heritabilities and breeding values. Microsatellites are preferred molecular markers for individual identification and parentage verification. Present study aimed at verifying parentage in daughters covered under progeny testing program operated by Dudhsagar Research and Development Association (DURDA), Mehsana. A total number of 212 Mehsana buffalo samples including 100 daughters for parentage testing, 12 sires and 100 dams were genotyped. Multiplex primer panel containing set of eleven fluorescent labeled microsatellite markers (CSSM61, ILSTS29, ILSTS17, ILSTS28, CSSM43, CSSM57, CSSM22, ILSTS61, CSSM8, ETH152, ILSTSl 1) was developed after screening of 22 microsatellites selected fi-om available list of 25 microsatellites suggested by the National Bureau of Animal Genetic Resources (NBAGR) and 25 cattle specific microsatellites. Primers were selected after their successful amplification individually and in multiplex panel. Multiplex panel was designed in a way that amplification of any primer does not overiap with amplification of other primer with the same dye. Multiplex panel was standardized and optimized for primer concentration and annealing temperature. Eleven microsatellites were amplified in single multiplex for automated fluorescence genotyping to verify parentage in Mehsana buffalo breeds. The number of alleles varied from 5 for marker ILSTSll to 16 for marker ILSTS61. The heterozygosity of the 11 different markers ranged between 0.281 for marker CSSM43 and 0.821 for marker ILSTS61. Calculated PIC values ranged from 0.80 highest for marker CSSM61 to 0.60 lowest for marker ILSTS29. Mean number of alleles per locus (k) was 9.91, Mean observed heterozygosity (HObs) was 0.675, Mean expected heterozygosity (HExp) was 0.762, Mean PIC observed was 0.730. Exclusion probability was highest for marker ILSTS61 and CSSM 61(0.49) and lowest for marker ILSTS29 and CSSM57 (0.25). The combined exclusion probability of all 10 markers was 0.993. Combined exclusion probability using five most polymorphic markers was 95.5 % and for all ten markers was 99.3 %. Paternity of 19 of 100 daughters (19%) was excluded because putative paternal alleles were not present in the progeny for at least two locus. Parentage was assigned to 95 daughters with strict confidence using Cervus 3.0 and five daughters were declared unassigned. Parentage verification also performed manually to check the results obtained by Cervus and they were in accordance. The developed panel of 10 microsatellites in a single multiplex constituted a fast, robust, reliable and economic tool to verify the parentage and assign the putative sire to daughters under progeny testing with very high accuracy. This can be used in routine parentage testing.
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ANIMAL BIOTECHNOLOGY, AN ANALYSIS