Morphological,chromosome Constitution And Gene Expression Studies Of Two Brassica Auto-allohexaploid(C~cC~cC~oC~oB~cB~c)

Polyploidization is a common phenomenon in nature and an important driving force for the plant speciation.The process of plant polyploidization is often accompanied by genetic,epigenetic and gene expression changes,resulting in phenotypes that are different from those of the parent,and thus adapt to the new environment.In this study,Brassica auto-allohexaploid(C~cC~cC~oC~oB~cB~c)was synthesized by the wide hybridization between B.carinata(2n=34,B~cB~cC~cC~c)and B.oleracea(C~oC~o,2n=18).Two young plants were differentiated from the callus of the same immature embryo during the embryo rescue on MS medium with colchicine and hormone.However,two plants have different phenotypes,one of which is purple in leaf and stem like the maternal parent B.carinata,while the other is green like the paternal parent B.oleracea.Cytogenetic,whole genome resequencing and transcriptome analysis were performed to find out the potential cause for the different phenotype of two plants.The main results are described as follows:1. Auto-allohexaploid synthesis and phenotype observationTwo types of plants(purple and green plant)were obtained by wide hybridization and chromosome doubling between B.cariana(G0-7)and B.oleracea(Chijielan).Purple-red color appeared in all tissues and organs except for the petals of purple plant,especially in young leaves,young flower buds and siliques,but no purple color was observed in green plant.In addition,the petals of purple plants are light yellow when they are just opened,and then gradually turn white,but the petals of green plants are white when they are opened.2. Chromosome number and composition analysis of two auto-allohexaploidsThe results of cytology and in situ hybridization showed that both green and purple plants had the expected 52 chromosomes including 16 chromosomes from B genome and36 chromosomes from C genome.The analysis of B-genome chromosome specific SSR markers showed that the chromosome B04 of the green plant was missing or partially missing.The further analysis of 20 SSR markers covering B04 chromosome showed that there was partial deletion of B04 chromosome.Whole-genome resequencing and reads mapping analysis confirmed that both the purple and green plants had 16 B genome chromosomes and 36 C genome chromosomes,but B04 chromosome of green plants lost one chromosome arm(0-22.7 MB).3. Comparative leave transcriptome analysis of two auto-allohexaploid leavesThe results of leaf transcriptome analysis showed that the average gene expression of B04 was significantly lower.A total of 7344 differentially expressed genes(DEGs)were identified between purple and green plants.Among 967 genes located in the missing part of B04 chromosome,965 genes were down regulated in green plants.There are 38 genes related to anthocyanin synthesis in DEGs,the expressions of which are all significantly lower in green plant than in purple plant,and only DFR(Bjub001305)gene is in the deletion region of B04 chromosome.Among the six genes related to carotenoid synthesis,two genes which located in deletion region of B04 chromosome,CRTISO(Bjub028062)and Z-ISO(Bju B042707)were significantly down-regulated in green plants.4. Comparative leave transcriptome analysis of two auto-allohexaploid petalsA total of 4028 differentially expressed genes(DEGs)were identified in petals between purple and green plants.Among 1212 genes located in the missing part of B04chromosome,1209 genes were down regulated in green plants.There are 13 genes related to anthocyanin synthesis in DEGs,but no gene in the deletion region.Among the seven genes related to carotenoid synthesis,two genes as revealed in leaves in deletion region of B04 chromosome,CRTISO(Bjub028062)and Z-ISO(Bju B042707)were significantly down-regulated in green plants.In conclusion,a B04 chromosome arm deletion mutant of Brassica auto-allohexaploid was identified by cytological,molecular marker and whole-genome resequencing analysis.The key gene for anthocyanin and carotenoid synthesis was primarily identified by transcriptome analysis combined with gene information in deletion region.The results will lay a foundation for the functional analysis of homologous genes of Brassica polyploid in anthocyanins and carotenoid synthesis pathway.