| The Anaka burmensis Dworakowska belongs to Hemiptera,Cicadellidae,Typhlocybinae,is the most important pest of bamboo leafhopper.There are many species of bamboo leafhopper,which are similar in appearance,so it is difficult to identify species rapidly and ac curately by traditional taxonomy.In order to understand the genetic structure of Anaka burmensis in different population,the mitochondrial DNA COI,16 S rRNA,and Cyt b gene sequence of leafhopper from 26 geological populations were analyzed to determine the genetic diversity,and analysis of the genetic differentiation,to explore the genetic factors of this insect.The results of the study are useful for understanding of the insect distribution and changes in Chinese,and for the molecular biology in the leafhopper insects,which can provide a theoretical basis for future research in molecular biology and the control of this insect.The major research results are described as follows:1.Analys is of genetic diversity among different geographic of Anaka burmensis based on mitochondrial COI gene sequencesMitochondrial DNA COI gene sequences of 241 individual leafhoppers from 26 different geographical population were cloned and sequenced.We calculated nucleotide composition,the genetic distance among the populations,the molecular variation analys is.The median-joining network was constructed by using Network 5000.Through the sequential analysis,we found that the value of A+T was about 72.4%,546 conserved sites,69 variable sites and 33 haplotypes were substituted in the 615 bp fragment of leafhoppers.The average genetic distance is 0.0166.Haplotype diversity index(Hd)and mean nucleotide diversity index(Pi)were 0.845 and 0.00877 respectively.The differences of the Neutral test Tajima 's D is-1.65898,Fu' s FS is-5.787,and the result is not signif icant(0.05< P <0.10).The coefficient of genetic differentiation among populations(Fst)and gene flow(Nst)were 0.72915 and 0.5985.The analysis of molecular variance showed that a high proportion of the total genetic variance was attributable to var iations among populations(72.92%).According to the constructed haplotype network map,the population of all round regions showed a phenomenon of high frequency haplotype spreading to low frequency.2.Analys is of genetic diversity among different geographic of Anaka burmensis based on mitochondrial 16 S rRNA gene sequencesMitochondrial DNA 16 S rRNA gene sequences of 243 individual leafhoppers from 26 different geographical population were cloned and sequenced.We calculated nucleotide composition,the genetic distance among the populations,the molecular variation analysis.Through the sequential analysis,we found that the value of A+T was about 78.7%,442 conserved sites,29 variable sites and 19 haplotypes w ere substituted in the 471 bp fragment of leafhoppers.The average genetic distance is 0.0020.Haplotype diversity index(Hd)and mean nucleotide diversity index(Pi)respectively were 0.332 and 0.00198.The differences of the Neutral test Tajima 's D is-2.22989,Fu' s FS is-14.788,and the result is very signif icantly different(P<0.10).The coefficient of genetic differentiation among populations(Fst)and gene flow(Nst)were 0.2654 and 0.2656.The analysis of molecular variance showed that a high proportion of the total genetic variance was attributable to variations among populations(63.23%).3.Analys is of genetic diversity among different geographic of Anaka burmensis based on mitochondrial Cyt b gene sequencesMitochondrial DNA Cyt b gene sequences of 200 individual leafhoppers from 22 different geographical population were cloned and sequenced.We calculated nucleotide composition,the genetic distance among the populations,the molecular variation analysis.Through the sequential analys is,we found that the value of A+T was about 78.1%,546 conserved sites,47 variable sites and 28 haplotypes were substituted in the 593 bp fragment of leafhoppers.The average genetic distance is 0.0057.Haplotype diversity index(Hd)and mean nucleotide diversity index(Pi)respectively were 0.830 and 0.00562.The differences of the Neutral test Tajima 's D is-1.80834,Fu' s FS is-9.171,and the result is signif icantly different(P<0.05).The coefficient of genetic differentiation among populations(Fst)and gene flow(Nst)were 0.6727 and 0.6729.The analysis of molecular variance showed that a high proportion of the total genetic variance was attributable to variations among populations(77.33%).In conclusion,there is no direct relationship between the genetic distance and geographical distribution of different geographical populations in Anaka burmensis based on the mitochondrial gene sequence,but there are significant differences among different geographical populations.4.The relationship between Anaka burmensis and approximate speciesUsing the three mitochondrial genes COI,16 S,rRNA,Cyt b,we explored the relationship between Anaka burmensis and its approximate species.,and constructed ML and NJ trees.The results showed that the genetic distances of three mitochondrial sequences in Anaka burmensis and Anaka sp.were 0.1528~0.1731?0.1459~0.2283?0.1779~0.1924;and the genetic distance between Sweta bambusana were 0.2101~0.3213?0.1608~0.1668?0.2547~0.3067;and the genetic distance between Trifida quadripunctata were 0.2437~0.2635?0.2020~0.2524?0.2114~0.2375.The genetic distance of Anaka burmensis within 0.0166~0.0490 can be determined to species;Interspecific genetic distances at 0.1528~0.2547 could identify different species.The NJ and ML tree suggests that the geographical population patterns of Anaka burmensis are in a parallel relationship.The topology of the tree is divided into two branches.The genetic distance between Anaka burmensis and Sweta bambusana is near than Trif ida quadripunctata.The genetic distance between Anaka burmensis and Anaka sp.is nearest.The gene Cyt b was screened as a molecular marker to identify different species,and a molecular system was preliminarily established. |
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