Genetic Diversity Of Two Major Brassica Crops And Utilization Of A Herbicide-Resistant Rapeseed Mutant In Breeding

Rapeseed is the most important oil crop in China.It has a strong heterosis.Chemical hybridization agents（CHAs）is one of the important approaches for utilization of heterosis in rapeseed.When CHA-resistant materials（the main active ingredient is herbicide）used as the male parent,it does not need to be protected with a shield or barrier,which would simplythe seed production protocol.The corresponding herbicides can be used to control weeds for herbicide-resistant rapeseed varieties in production.Therefore,the breeding ofherbicide-resistantrapeseedis of great significance.Genetic diversity analysisis an important basis work for germplasm conservation and breeding.Previously,our labhas obtained a TBM-resistant mutant K5 in M₂population of the rapeseed cultivar ZS09 treated by ethylmethanesulfonate（EMS）.In the present study,the homozygous mutant line derived from K5 wasused as material,and the following studies were carried out:1)A set of 100 Brassicaaccessions（mainly including Brassica napusand B.juncea）were evaluated for their genetic diversity bysimple sequence repeats（SSR）,sequence-related amplified polymorphism（SRAP）and intron-exon splice junctions（ISJ）molecular markers.Furthermore,the cytoplasmic type of the tested accessionswas identified by a multiplex PCR assay method.The results obtained will be significant for germplasm conservation and breeding in rapeseed;2)Effects of CHA on floral organ morphology,pollen fertility,and yield in different rapeseed accessions.The results are as follows:1.Aset of 100 Brassicaaccessions,including 50 B.juncea,31 B.napus,fiveB.rapa,fiveB.carinata,fourB.oleracea,threeB.nigra,one Eruca sativaand oneIsatistinctoria,was genotyped by SSR,SRAP and ISJ markers.The results showed that a total of 87 polymorphic bands were detected by 23 pairs of SSR primers,each pair of primer could amplify 1-6polymorphic bands,and the average polymorphic information content（PIC）value was 0.64.Two hundred and eighteen polymorphic fragments were amplified by using 27 pairs of SRAP primers,the average PIC value was 0.85.Seventy two polymorphic bands were detected by eight ISJ primers,the number of polymorphic bands amplified ranged from six to 13,the average PIC value is 0.81.A total of 377 polymorphic markers（SSR,SRAP and ISJ markers）was used to perform cluster analysis of the 100 Brassica accessions.The results showed that the tested lines were divided into 6groups.All 31 B.napusand 50 B.juncea were grouped together,respectively.Other materials were also grouped together according to their types.The results of principal component analysis andpopulation structure analysis were basically consistent with the results of cluster analysis.Cytoplasmic type identification of the 100 tested materials was performed by using one-step multiplex PCR assay method.The results showed that among 50 B.juncea,49 were Cam type and onewas Nap type,among 31 B.napus,11 were Pol type,threeCam type,15 Nap type,and two mixed types.These results indicated that the cytoplasmic type of B.juncea was simple,whereas that of B.napuswas relatively richer.2.The B.napus mutant K5 and 12 elite lines were used as materials,0.1%CHA-XN01was applied toproduce hybrid seed.As a result,the 12 accessions showed complete male sterility after treated with CHA,they exhibited short filaments,dried stamens and inactivated pollens.However,the K5 showed normalflower organ and pollen viabilityas the control plant ZS09 without CHA treatment.Seed yield of K5 was 109.72 Kg/667m²,which was the highest in the testedaccessions,indicating that the CHA had the least impact on seed yield of K5.The seed yield of the 12 lines ranged from 21.01 to 83.65 kg/667 m².