Genetic Polymorphism Analysis Of Mtdna D-Loop Region In Different Populations Of The Tilapia(Oreochromis)

In this study,159samples of6tilapia populations were collected from Egyptian Oreochromis niloticus (AN、GN,22、28samples)、88strain O. niloticus (XN,26samples)、 American O. niloticus (MN,26samples) and O. aureus (ALY、GA,28、29samples). We extracted the gene group DNA using the method of phenol/chloroform. The partial sequences of159individuals of6tilapia populations were amplified by PCR and sequenced. Then we aligned and edited all these sequence using Clustal W, Bioedit, EditSeq software. We also statisticed the variation site、genetic distance and constructed the NJ and UPGMA tree by the software of MEGA3.1. Nucleotide polymorphism, haplotype and gene flow among populations were statisticed by the software of DNASP4. The results are presented as follows:1. Tilapia partial sequences of mtDNA D-loop were575-581bp.111sites were polymorphism, which were19.2%of the whole sequence, with47singleton polymorphic sites and57parsimony informative polymorphic sites. The three bases singleton polymorphic sites were not found. There were five kinds of mutation of nucleotide. They were transition, transversion, insertion/deletion and the coexistence of transition and transversion.80sites were transition and24sites were transversion in104replacement sites. The ratio of transition and tranversion was3.3. There were four insert/deletion sites (234,253,360,419), one insertion site (380), two deletion sites (33,162). The ratio of base replacement and insertion/deletion was14.9. The nucleotide replacement existed mainly in transition (T(?)C or A(?)G).2. In159mtDNA D-loop sequences of six tilapia populations, averaged nucleotide content were:28.8%for A,15.4%for G,35.4%for T and20.4%for C. Averaged A+T content(64.2%) was excess of G+C(35.8%) in tilapia D-loop region. The tilapia mtDNA D-loop region was rich in A+T, which had highly base bias. The central conserved sequence block and conserved sequence block of mtDNA D-loop region were identified. The four conserved sequence blocks (CSB-D、CSB-1、CSB-2、CSB-3) were successfully identified and their homology compared with other fish. 3. Comparison of these159mtDNA D-loop sequences revealed52mitochondrial haplotypes in tilapia. The number of haplotypes was12for the AN population,12for GN population,10for the XN population,11for MN population,9for the ALY population and10for GA population, respectively. One haplotype sequence was shared by six populations. One haplotype was shared by AN、GN and MN populations; one haplotype was shared by XN, MN and GN populations too. One haplotype was shared by AN and XN, so was in AN and GN, MN and GN. Others were special in each population. It demonstrated that tilapia presented certain mitochondrial genetic diversity.4. According to the gene flow within populations, there was no divergence between AN and GN、ALY and GA. But there was divergence among other populations. From genetic distance, the D value between AN and ALY was largest and the distance between MN and ALY is minimum. It could find that the hyprid offspring of AN and ALY had better hybrid advantages, which had the foundation of further selective breeding.5. According to the genetic diversity parameter nucleotide polymorphism (Pi), the value of O. niloticus was higher than O. aureus. The NJ tree was divided into two parts, in which ALY, GA were together, but AN、GN、XN and MN were mixed. It manifested the purity of ALY was high; the genetic diversity of AN and XH was rich and trait stable.