Study On Polymorphism Of Mitochondrial DNA From Portunus Trituberculatus

This experement is study on the population distribution of Portunus trituberculatus along China shores.The samples come from Dandong,Yingkou,Laizhou, Qingdao, Lianyungang, Ningbo, Beihai.And the sample is wild individuals.Study on partical sequence of mtDNA 16S rRNA Cytb and D-Loop,we can research the genetic structure and genetic diversity of Portunus trituberculatus in the China sea. All these results present a basis to the conservation and utilization of genetic diversity of Portunus trituberculatus.The genomic DNA sequnence was extracted by high salts method. Applying Primer Premier 5.0 to design PCR primer and selecte of polymorphic primers to three mitochondrial DNA gene fragments: 16S rRNA Cytb and D-Loop.Then the results of PCR were sequenced.Alignment of the results of PCR was done by Clustal X 1.83 package.Evolutionary distances were calculated by using Kuimura 2-Parameter distance model of MEGA software package version 3.1 and phylogenetic tree was contstructed by Neighber-Jioning Method.1000 samples were chosen randomly to caculate the bootstrap value for evalauating the confidence level of phylogenetic tree. Haplotype diversity (h)(Nei &Tajima,1981)and nucleotide diversity (Ï€)(Nei,1987)were calculated by using the DnaSP software Version 3.53 to evaluate the genetic diversity in 7 populations of Portunus trituberculatus.In order to reflect the genetic differentiation between and among populations, analysis of molecular variance (AMOVA) (Excoffier et al.1992)was done by Arlequin software package version 2.0(Schneider et al.2000).1.A 514 bp length of 16s rRNA gene fragment was obtained.there was no different among individuals and there was no difference between individuals.The phylogenetic tree based on Portunus trituberculatus and 42 related species was set up by NJ method. The result of cluster is consistent with traditional morphological classification.2.A 644 bp length of Cytb gene fragment was obtained, only two transformation sites.3. A 617bp length of DNA fragment was obtained .Sequnence analysis showed the sequence of the 48 individuals with each other ,80 variation sites was observed,of which there 6 insertion sites,22 transition/transversion sites. The range herterozygosity value of local population is 0.030~0.014.The genetic distance between the individuals is 0.021 on the average.The genetic diversity of D-Loop genome betweene each population is very small,and which belows the genetic diversity of subfamliy. The greatest genetic diatance(0.009)occurred among the LaiZhou population And very small genetic coefficients of variation for genetic variation.The most genetic distance occurred among the LianYunGang population,Seconed is the NingBo population,the next is the QingDao population.The reason is perhaps to be found in the locations of Various Species.4. A high level of genetic different was detected among populations with analysis of molecular variance. Analysis of molecular variance (AMOVA)showed that there was 17.77% of variant component among populations,and 82.23 % of variant compontent within populations.5. 38 haplotypes in 48 samples,and it contain 9 shared haplotypes.4 of 9 shared haplotypes shared between different region ,5 of 9 shared haplotypes shared between the same region. The LaiZhou population and the BeiHai populationHave their own maternal origin,and they become their geographical population, Resources development the two endemic species were suggested.