Molecular Cloning And Expression Analysis Of DsGPAT、DsPDAT Genes From Descurainia Sophia

Descurainia Sophia, a cruciferous annual herb species, has relatively strong ecological adaptability, and contains abundant unsaturated fatty acids in seeds. And it is of great significance to research D.sophia gene resources. Objectives of present paper are to clone and characterize the two genes related to lipid synthesis from D. Sophia DsGPAT、DsPDAT, for further improving oil contant of Brassica napus. The main results are as follows:1. Cloning of two important genes related to seed lipid accumulationGlycerol-3-phosphate acyltransferase (GPAT) involved in the first acylation in the TAG biosynthesis was demonstrated to be important to lipid accumulation. cDNAs of DsPDAT was isolated from D.sophia using RACE-PCR. The full-length of DsGPAT cDNA encods 462 amino acids with molecular weight 53.53 KDa, The isoelectric point is 6.74. By software analysis, the DsGPAT is predicted to contain a chloroplast signal peptide at N-terminus that may target to chloroplast. DsGPAT do not contain transmembrane domain, but it has the highly conserved functional domains between 226-372 which is the characteristic of the acyltransferase.The phospholipid:diacylglycerol acyl transferase (PDAT) participates in the synthesis process of DAG to TAG, and may be very important in seed lipid accumulation. cDNA length of DsPDAT is 2227 bp, ORF is 1995 bp, encoding 664 amino acids with molecular weight 44.1 KD, isoelectric point 8.63. DsGPAT protein with N-terminal hydrophobic has a very typical transmembrane domain at the 42-64. It has a highly conserved domain-LACT between the 114-627 which is typical structure shared by LACT family members.2. Expression analysis of the two genesThe stems, leaves, bud, flower, silique of D. Sophia were harvested for quantitative RT-PCR detection with 18S and 5.8SrRNA as internal reference The results showed that the two genes are constitutively expressed in these tissues. The expression of DsPDAT and DsGPAThave similar trend in the bud, flower and silique. Expression for the two genes were both gradually increased from the lowest in leaves to highest in siliques, indicating that the two genes are probably involved in the process of lipid accumulation.