The Study Of Determining The Relationship In Eleven Kinds Of The Longicorn Beetles By Rapd Molecular Mark And Its-2 Sequence

This thesis has explored several kinds of methods for extract DNA template rapidly and conveniently from dry, liquid-dipped or fresh specimens of longicorn beetles, so the most efficient methods were gotten for 3 kinds of specimens under different environment respectively. The results of PCR analysis to the genome DNA that has already got indicated these extraction methods were feasible. Carry on research to 11 kinds of longicorn beetles by two molecular marking techniques (RAPD, ITS-2). The results :1. In general, the difference of Coleoptera insects from others is that the body wall is very hard, gas and water permeability is bad. Under the same conditions the time for drying is longer and the DNA quality worse than other kinds. In order to get the high quality DNA longicorn beetles had better be dried or icy. The genomic DNA of the museum specimens kept for a long time degradated very seriously and had the factors for influencing PCR. The primary treatment of extracting the genomic DNA from museum specimens is that the specimens were dipped into the STE buffer for 24 hours, then washed three times with deionizing water. This method can greatly reduce the rupture of the genomic DNA and be in favor of protecting the genomic DNA in the museum specimens, and abate or eliminate the influence on PCR.2. The RAPD-PCR figures showed DNA diversity is obvious, and PCR electrophoretograms of one kind insect under the different stage (larvae and adult) were same. It is feasible for the DNA diversity of related species and identification of important species.3. PAPD analysis of 11 kinds of longicorn beetles genetic relationship was carried out with 16 random primers. The study had detected 157 loci. The percentage of polymorphic loci shares a range of 5.10%~12.10%. X. rusticus (12.10%) is the highest and S. bifasciatus (5.10%) is the lowest. Genetic relationship cluster figure of 11 kinds of longicorn beetles was got by RAPD marking technique.4. ITS-2 sequences of 9 kinds of longicorns longicorn beetles has already been received by Genbank. Further analysis on relationship among 9 kinds of longicorn beetles through the ITS-2 sequences of rDNA was carried out, the length of ITS-2 sequence ranged from 324bp to 381bp. Full length sequences of ITS-2 were obtained after sequencing, and the sequence data were used to construct the phylogenetic tree of 9 longicorns species.Keywords Longicorn beetle; genomic DNA extraction; RAPD; ITS-2; relationship...