Genome-wide Identification And Functional Analysis Of CLE Gene Family In Brassica Napus L

CLE(CLAVATA3/Embryo surrounding region-related)gene family encodes a signaling peptide and plays a key role in the cell – cell communication,cell proliferation and differentiation,self-incompatibility determination and defense response.Several researches have been carried out on CLE gene family from various crops such as Arabidopsis thaliana,Oryza sativa,Glycine max,Solanum lycopersicum and Populus tremula e.t.c,but there is a dearth of information about the functionality of CLE polypeptide genes in the Brassica family.In this study,bioinformatics and in vitro peptides application were used to analyze the functions of CLE gene family in Brassica napus,while the expression pattern of these genes were measured using qRT-PCR.The results obtained are as follows:1.116 CLE peptide-encoding genes were identified in Brassica napus genome.These identified genes contain seven new genes and five gene protein sequences with multiple CLE domain.In addition to that,the signal peptide prediction,isoelectric point,molecular weight,polypeptide sequence length and gene structure were characterized.2.Chromosome localization revealed that the chromosomes A and C contain 53 and 61 BnCLE genes respectively.It was observed that BnCLE genes always occurs in pairs in the homologous region of the chromosome.Segmental duplication is the main expansion way of BnCLE gene family(68%,79/116)when compared to whole-genome duplication which accounts for 27% and 31/116,while the least rate of expansion(5%,6/116)was identified to be tandemly duplicated.3.Full length protein sequences were used for the multiple sequence alignment and phylogenetic tree.According to the AtCLE genes functional classification in Arabidopsis,BnCLE genes were classified into three categories according to their functions.Group A mainly acts on the growth and development of lateral root while group B mainly maintain the stem cells dynamic balance in stem and root apical meristems.Group C members play a key role in the development of vascular cambium.4.Five BnCLE genes with multi-CLE domains were identified.It revealed that the five BnCLE genes with multi-CLE domains are highly expressed in ovary and silique.This shows that these genes may play a key role in the development of seeds.5.45 BnCLE genes containing 29 types of new CLE motif were identified.In vitro polypeptide processing was used to verify the function of 27 out of the 29 CLE peptides.The results showed that most of the distinct CLE peptides in B.napus were homologous to A.thaliana and still maintain consistent functions while there are few occurrences of functional differentiation.6.Ascertaining the expression level of 48 genes in the B.napus tissue showed that the expression level of most BnCLE genes are tissue specific.The expression differentiation of some homologous BnCLE genes confirmed that the functional sites were differentiated.The above results laid a theoretical foundation for the CLE gene function and specific identification in rapeseed.