Studies On Analysis Of Heredity Regular And RAPD Markers Of Gynoecious In Muskmelon (Cucumis Melo L.)

The inheritance of gynoecious was researched according to segregation of every populations in melon (Cucumis melo L.).Bulked segregant and RAPD analyses were employed to identify molecular linked to gynoecious character. The possibility of Markers assisted selection was explored in gynoecious selfing line which could speed up seed breeding and develop a new method in melon. Below are the details in this research.Two F1,two BC1 and two F2 populations, were obtained from the two crossed between gynoecious and Hetao-Mi, and gynoecious and Lv-Bing. Segregation of every generation were investigated and confirmed the gene type of gynoecious was A_gggygy, and the gy was modificatory gene.The CTAB method was employed to isolate genome DNA. The CTAB method was time consuming, with much more producers. And the quality of extracted DNA was high and was fully digested by restriction endoenzyme, EcoR I. DNA samples isolated by CTAB method was fit for PCR.The main factors effecting RAPD analyses were discussed and the conditions of the RAPD reaction were optimized, including optimum template, dNTP, primer concentrations, and PCR proceducers. A reliable and reproducible RAPD protocol was obtained in this research.400 decamer primers were used to screen two pairs of bulks. Bulk1 and bulk3 were gynoecious DNA pool; Bulk2 and bulk4 were andromonoecious. 93.5% of the primers used in these experiments produced distinct amplified products when both gynoecious and andromonoecious bulks were used. Totally, there were 2396 bands amplified by 374 primers in bulk1 and bulk2, and 2468 bands in bulk2 and bulk4. The average amplified bands per primer were 6.4 and 6.6 respectively. 15 primers had polymorphic bands between bulk1 and bulk2, while 11 primers had polymorphic bands between bulk3 and bulk4. In addition, 4 primers had polymorphic bands between two pairs of bulks. After RAPD analyses of 1 individuals selected randomly in each bulks, only S1001 amplified a 920bp polymorphic band in all individuals from bulk1 and bulk3 which was absent in all individuals from bulk2 and bulk4. This polymorphic band named S1008920.These specific DNA bands amplified in bulk1 and bulk3 were recovered, and then inserted into pGEM-T Easy Vector plasmid, and finally transported into E coli. DH5α.Recombinant plasmid. DNA was isolated from white plaques of E coli. After digested by EcoRⅠ, it was proved that the polymorphic fragments S1008920 were transported into E coli. successfully. According to the sequence of S1008920, a pair of 22-mer specific primers were designed. After PCR using this pair of primers, the specific band S1008920 had no polymorphic in both two F2 populations. And the factors which effected in transferring RAPD markers into SCAR markers were discussed.