Preliminary Analysis Of Cloning And Expression Characteristics Of Sugar Beet Chlorophyll Ab Binding Protein Gene BvLhcb-2

Sugar beet(Beta vulgaris L.)is an important sugar crop.In order to promote the development of sugar beet industry,in addition to some conventional cultivation and breeding methods,trait improvement can also be realized at the molecular level through gene operation.However,sugar beet gene resources are not abundant in China at present,so how to obtain enough gene resources has become an important topic in the research of sugar beet breeding.Based on the previous work of the research group,this study analyzed the differentially expressed transcripts in sugar beet germplasm with different leaf colors,the beet chlorophyll a-b binding protein gene were picked out as the target gene,named BvLhcb-2,both the coding region fragment of the target gene and its complete gene locus fragment in the genome were isolated by nested PCR.Then the exact sequence of the target gene was analyzed on its bioinformatic and expression characteristics.The effects of drought,weak light and aging conditions on the expression of BvLhcb-2 were analyzed.This study will enrich sugar beet gene resources.The results can provide some information on leaf color regulation and photosynthesis,and provide relevant theoretical basis for subsequent scientific research and application.The main results are as follows:(1)The BvLhcb-2 gene of Sugarbeet was isolated and its sequence integrity was verified.Through a series of steps such as primer design,nested PCR amplification,cloning vector construction and sequencing,an 804 bp sequence was obtained.The integrity of the sequence was determined by ORF finder and blast X online software.The sequence structure of the target gene in the genome is the same as that in m RNA,and there is no intron in the coding region.(2)Bioinformatics analysis of BvLhcb-2 gene and its encoding amino acid sequence was carried out.By analyzing BvLhcb-2 gene,it was found that it had high similarity with sugar beet chlorophyll A-B binding protein gene.Through the analysis of BvLhcb-2 protein,it is inferred that the protein belongs to chlorophyll A-B binding superfamily.The length of the coding product is 267 aa,the molecular weight is28286.31,and the isoelectric point is 5.16.It has no transmembrane region,hydrophilic,no signal peptide,and has a total of 18 phosphorylation sites.(3)The expression characteristics of the target gene were analyzed.Four groups of treatments were set up: drought,shading,abscisic acid and ethylene.Firstly,the changes of external morphological characteristics and chlorophyll content of sugar beet under different treatments were analyzed through preliminary experiments,and then the expression patterns of BvLhcb-2 gene in different sugar beet germplasms under different treatments were analyzed by Quantitative Real-time PCR.The results showed that under different treatments,the variation trends of genes in the two germplasm were different.Under drought treatment,the expression of BvLhcb-2 first increased and then decreased.After rehydration,the expression of BvLhcb-2 was down-regulated in 'J9451',but significantly up-regulated in 'DZ-6';Under the shading treatment,the gene first decreased and then increased in 'J9451',but it first increased and then decreased in'DZ-6';Under abscisic acid and ethylene treatment,the gene showed an overall downward trend in 'J9451',while in 'DZ-6' showed a trend of first rising and then falling.