Screening Microsatellite Markers For The Yangtze Finless Porpoise And Applying In Parentage Identification

The Yangtze finless porpoise (Neophocaena phocaenoides asiaeorientalis) is known to be an unique freshwater subspecies of Neophocaena phocaenoides, which distributes in the middle and lower reaches of the Yangtze River and its appended Dongting and Poyang Lake. In recent years, expanding human activities in the Yangtze River has caused the habitat of the Yangtze finless porpoise to deteriorate, resulted in dramatically drop in population size. The most recent ecological survey indicated that only about 1800 individuals left in the Yangtze River. As concerned its worsening population status in the wild, it was listed as an endangered population in the IUCN Red Data Book, and as second order key protected animals by Chinese government. Meanwhile, it is very urgent to strengthen its conservation strategies and scientific research as well.For the Yangtze finless porpoise, an ex situ conservation breeding population of about 30 individuals has been successfully established at Tian-e-Zhou National Natural Reserve with 2 to 4 young born each year, besides, a small captivity-bred colony was also raised in the Baiji Dolphinarium that gave birth to three calves in the past 3 years. However, scientific and effective genetic managements based on proper technique of parentage identification are required to the final success of all ex situ conservation endeavor for the Yangtze finless porpoise. Till now, microsatellite parentage identification technique has been successfully applied in the protection and management of many rare and endangered species. To the aim of establishing a similar technique for parentage identification in the Yangtze finless porpoise, polymorphic microsatellite loci have been rapidly and successfully screened through ways of PCR amplification and non-denatured PAGE electrophoresis. And then, 8 polymorphic loci were selected to genotype three Yangtze finless porpoise populations with fluorescence labeling primers. Finally, for the two small stocks that raised by Wuhan Baiji Dolphinarium and Tongling National Natural Reserve, all of the parentage relationships have been successfully identified.Main results obtained in this dissertation were summarized as follows: (1) Fourteen specific microsatellite primer pairs were newly designed for the Yangtze finless porpoise, and 11 of them were proved to be polymorphic through PCR amplification and non-denatured PAGE electrophoresis.(2) Polymorphism of 8 specific microsatellite loci in the finless porpoise was analyzed by genotyping after PCR amplification with fluorescent dye 6-FAM labeled primers. As results, for either of the three Yangtze finless porpoise populations explored in this study, all of the 8 loci were proved to be polymorphic with more than 2 alleles at each locus. Besides, 6 of them shown to be highly polymorphic loci with polymorphism information content (PIC) values larger than 0.5.(3) Parentage identifications were conducted among two small Yangtze finless porpoise breeding stocks that translocated and reared by Wuhan Baiji Dolphinarium and Tongling National Natural Reserve respectively. Because the mothers were known definitely, the fathers for all of the offspring born in the two stocks were easily and successfully identified by genotyping with 8 polymorphic microsatellite loci.In summary, after this dissertation, many more useful microsatellite genetic markers have been accumulated for conservation genetic study of the Yangtze finless porpoise. Since they were proved to be successful in parentage identification of the Yangtze finless porpoise in this study, they should provide a solid technical base for the establishment of studbooks and genetic management strategies on different ex situ conservative populations in the coming future.