Cloning And Expression Analysis Of StRALF Gene In Setosphaeria Turcica

The maize leaf blight is a major leaf disease in the main maize producing areas of the world,which has caused serious economic losses.It happened more frequently in the northeast,northwest,north of northern China and the mountain area of south of China.In this article,basing on the rapid alkalinization factor(RALF)as the research object,the conserved sequence of St RALF in Setosphaeria turcica was cloned.RT-q PCR technique was used to analyze the gene expression in different acid-base conditions and different periods of the infection.The full length of the c DNA sequence was obtained by RACE,and the encoding protein of the gene was analyzed by bioinformatics.The results are as follows:This paper cloned the conserved sequence of St RALF in Setosphaeria turcica with a length of 114 bp.Bioinformatics analysis of its coding protein showed that this sequence encodes 37 amino acids.It was predicted that molecular weight is 3.6 KD and the theoretical isoelectric point is 7.74.According to the comparative analysis of amino acid homology,RALF protein has homology with published RALF protein,but it is not completely conservative.The full length of c DNA sequence of St RALF was obtained by RACE technology,which is 279 bp.Bioinformatics analysis showed that it encodes 92 amino acids,called secreted proteins.This protein does not contain spiral regions,so it cannot perform metabolic regulation,recognition of molecules,membrane channels and other functions.The results of amino acid sequence analysis showed that the sequence of c DNA and the conserved sequence of cloning of the St RALF were obtained successfully.Compared with other fast alkalinization factors of amino acid homology sequences were similar to the c DNA of St RALF.This will build up a good basis for the further studies on the functions of St RALF proteinSt RALF in Setosphaeria turcica was related to infection of the maize leaf blight.The phenotypic analysis of Setosphaeria turcica under different acid-base culture medium showed that the mycelium growth ability was strong;Under different acid-base conditions,the sporulation was high when the alkalinity was weak and the pathogenicity high under weak alkaline conditions.The expression level of St RALF was up-regulated and then decreased from acid to alkali culture conditions.The expression level of St RALF was relatively higher under weakly alkaline conditions than that under weakly acidic conditions.In the interaction with the host,the expression of St RALF was gradually up-regulated,reaching the maximum at 96 h,which was 5 times the expression at 24 h,suggesting that St RALF was related to the infection of the maize leaf blight.Under different acid-base conditions,the St RALF expression reached the maximum after inoculation for 96 hours under the condition of p H was 7.5 and infection rate was the highest.Under the condition of p H was 3.5,the expression level was the minimum and the infection rate was the lowest.It was indicated that the weakly alkaline environment promotes the expression of St RALF,which is contribute to infection.