Genetic And Relationships Analysis In Some Species Of Cycadaceae In China By Rapd Markers

Cycadaceae is the most remote antique seed plant existing in the world. It has important value of science and economy. In this paper, the RAPD reaction system in Cycadaceae has been established and applied to discuss the relationships among 21 species. The results showed as follow:1. The genome DNA of Cycadaceae with high concentration and purity could be obtained with the improved method of CTAB, which is good enough for RAPD amplification.2. The optimized reaction system of RAPD was summed up base d on the experimental results in this study. The system has 20μl react ion volume, including super pure water 9.04μl, 10×Taq buffer 2.0μl, 2 5mmoI/L Mg2+1.6μl, 2.5mmol/L dNTP 1.6μl, 10μmol/L primers 0.6μl, 5U/μl Taq Polymerase 0.2μl, 10ng/μl template DNA 5μl.The optimal PCR amplification procedure inclues: Pre-denaturation at 94℃ for 5min; and 35 cycles of denaturation at 94℃ for 30 sec, annealing at 37℃ for 40sec, lengthening at 72℃ for llOsec; and then again lengthening at 72 ℃ for 7 min. The final product was kept at 4℃.3. The results of RAPD cluster showed that the similarity coefficients of all species are ranged from 0.2903 to 0.7016, changing dramaticaly, showed the genetic background of Cycas is very complex. The similarity coefficient of C.multifida and C.segmentifida is 0.7016, which is the biggest one in thatof all species, showed they have closest genetic relationship. The similarity coefficient of C.longlinensis and C.multifrondis was 0.2903, which is the smallest one in that of all species, showed they have farest genetic relationship. The similarity coefficient of different sex colony of C.diannanensis is 0.9098, which is the biggest one of all material, they cluster first of all, showed the closest genetic relationship of all material. The results of RAPD cluster reflected the genetic relationship of all species, showed RAPD is suit for inter-species genetic relationship analysis of Cycadaceae.Genetic relationship analysis by RAPD marker combining with mo rphology showed that Cmultifida and C.segmentifida have close geneticrelationship, Cmultifida should be a sub-species of C.segmentifida. Th e species that have far genetic relationships of each other listed as fol low: Cxilingensis, C.longlinensis, C.segmentifida, Cacuminatissima, C.mi cholitzii, Clongipetiolultty C.multifrondis, Clongiconifera, Cguizhouensis,Cszechuanensis, Cbrevipinnata, Cmiquelii, Cbalansae, Cshiwandasha nicay Cparvulus, Cfairylakea, Chainanensis, Ctaiwaniana, Guangdong Cycad, C.diannanensis, each of them should be an independent species.