Molecular Cloning And Expression Analysis Of B-class MADS-box Genes From Brassica Oleracea Var Acephala

Brassica Oleracea var acephala is a horticultural mutant of the Cruciferae Brassica species,which is widely used in landscape gardening because of various colors of leaves and shapes.The wild type Brassica Oleracea have bisexual flower,four petals arranged in cross and six stamens.Two kinds of mutant were fond in breeding work.One is petal-loss mutant,in which the number of petal arranges form zero to three,and the other one has more petals,in which the number of petal arranges from five to muti-wheel.This research mainly focuses on the analysis of function relationship and the expression pattern of temporal and spacial of B class MADS-box genes,so that we can lay a solid foundation for explaining the molecular mechanism of kale mutants and flower development.(1)The morphological structure of Brassica Oleracea flowers organsObserve the phenotypic differences of flower organs and the differences of the primordium development between wild type and mutants in Brassica Oleracea by using anatomical entity lens and scanning electron microscope.The results show that the primordium of mutants had changed during early development.(2)The full-length cDNA sequences cloning of B class MADS-box genes in Brassica OleraceaUse RACE and other technologies to clone the full-length cDNA of B class MADS-box genes in Brassica Oleracea's.Achieve five genes which are BroaAP3a(accession number:KC896403)?BroaAP3b(accession number:KC896404)?BroaPIa(accession number:KC967961)?BroaPIb(accession number:KC967962)?BroaPIc(accession number:KC967963).(3)The phylogenetic analysis of MADS-box genesDetermine the affinity of BroaAP3a,BroaAP3b,BroaPIa,BroaPIb,BroaPIc protein by builting a phylogenetic tree.Through the analysis of MICK structure,it suggest that BroaAP3a and BroaAP3b belong to euAP3/DEF which derived form paleoAP3/DEF.BroaPla,BroaPlb and BroaPIc are belong to the PI/GLO.By predicting the primary and secondary structure of BroaAP3a,BroaAP3b,BroaPIa,BroaPIb,BroaPIc,it found that all of them are hydrophobic basic proteins.(4)The expression patterns analysis of temporal and spacial of Brassica Oleracea' s B class MADS-box genes by using semi-quantitative and Realtime methodBy analysising MADS-box genes specific expression of temporal and spacial during the process of floral development,the expression of BroaAP3a?BroaAP3b genes was found extend to sepals in mutants.The role of BroaPIc gene in Brassica Oleace a mutant' s petal development is very important.