Cytoplasmic Type Identification And Analysis In CMS Germplasm Of Radish (Raphanus Sativus L.)

Radish (Raphanus sativus L.) was originated in China and for important Brassicaceae vegetables, which had significant heterosis. Male-sterile was that plant could't produce functional pollens, while the growth of pistil and nutrition was normal, which was an important way for using heterosis. Ogura cytoplasm was found first and was widely studied CMS type. Now Ogura cytoplasm had been widely used in commercial production of hybrids.However, CMS lines using for production were Ogura cytoplasm, which could easily lead to the production at the risk of irreparable damage. The technical system of cytoplasm identification of radish germplasm has not been established, the abortion mechanism of process of microsporogenesis need to be studied, the selection of appropriate maintainer was difficult, the system of maintainer screened by MAS was not established. We carried out the cytoplasm identification in different radish germplasm, the process of microspore abortion was observed and maintainer screened by MAS was studied. It was hoped that rapid cytoplasm identification of radish, reveals the mechanism of microspore abortion, maintainers screened rapidly and the efficient use of CMS lines, which were provide an important technical foundation.(1) 231 inbred lines, CMS lines and 48 cultivars were amplified by specific primers designed based on atp6 and orfl38 genes, radish cytoplasm were divided into four types: DBRMF1, DBRMF2, Chiasmatype and Ogura.55 inbred lines were DBRMF1, only amplified 835bp fragment; 23 cultivars were DBRMF2, only amplified 910 fragment; 62 accessions were Chiasmatype, which could amplified both orf138 fragments and normal cytoplasmic fragments; Ogura type were 139, which could only amplified orf138 fragment. The orf138 play an important role in Ogura CMS, so Chiasmatype and Ogura type were Ogura cytoplasm. Radish cytoplasm was identified and classified by specific molecular marker, which provide an important technical foundation for the rational utilization and evaluation of germplasm resources.(2) Microsporogenesis of cytological was observed in three type accessions. It was showed that:the different CMS lines with the same paternal had similar way of abortion, and their microspores had consistent developmental condition before tetrad stage, Tapetal began disintegrating after the tetrad stage and microspores were aborted because of lack energy providing; The combination of fertility differentiation showed strong consistency, microspores showed differences after the tetrad stage, fertile accessions developed normal and form microspores, sterile accessions result in sterile because of tapetum squeezing microspore; there was a significant feature of tapetum squeezing microspore not thorough in half sterile accessions, there were still some microspores develop into mature pollen grains. This paper compared the process of microsporogenesis in different accessions, in order to determine the period of microspores and methods of abortion, and provide an important reference for cytoplasm identification.(3) 15 inbred lines and 17 CMS lines were amplified by primers designed based on orf138 and orfl58 genes. Specific bands of orf138 were amplified by primer Lw oguspecific 138FR in 21 accessions. Specific fragments of orfl58 were amplified by primer Lw 158FR in the all accessions. It was indicated that 21 accessions were the Ogura cytoplasm, restoring genes exist in the 3 inbred lines and couldn't used as the candidate maintainer; 11 inbred lines didn't including the orf138 specific band and suitable as a maintainer. Maintainers were hybrid with CMS lines, and eliminating the inbred lines which appeared to be fertility segregation, and combined with the performance of field trait, finaly breeding new CMS lines used for hybrid seed production in radish. Molecular markers based on field traits identification were used for the rapid screening of excellent maintainer, which reducing time and working of tesecross, increasing the efficiency of transfer and the process of CMS used in F1 cultivars production.