Cloning And Expression Profiling Analysis Of Soluble Starch Synthase Gene In Lotus(Nelumbo Mucifera Gaertn)

Lotus （Nelumbo nucifera Gaertn）, a member of the family Nymphaeaceae, is a perennial aquatic herb, which is wildly cultivated in china as a kinds of vegetable for a long history. Starch, composed of amylose and amylopectin is the main storage nutrition in lotus, and the content and ratio of amylose and amylopectin affect the quality of the products of lotus. Five types of important enzymes involved in the starch biosynthesis, such as:ADP-glucose pyrophospho-rylase polypeptide, granule-bound starch synthase, soluble starch synthase, starch branching enzyme and starch debranching enzyme. The soluble starch synthase is widely considered as one of the most important enzymes in plant amylopectin synthesis. Therefore, in this study, a full-length of cDNA and genomic DNA of SSS were isolated, and expression of this gene was also analyzed, we obtained following results.1. The immature leaves of lotus were chosen as material to extract the total RNA. A soluble starch synthase gene （SSS） was isolated using RACE-PCR technique. The full-length cDNA of this gene was 4080 bp, containing an open reading frame which was 3696 bp encoding 1231 amino acids. Compared this gene against NCBI database with nucleotide acid, SSS gene of’Mei ren hong’（KP201636） showed 99% similarity with’E lian 5’and’Sacred lotus’ （XP₀10278676.1）. In addition, thirteen differences in amino acid of SSS between’Mei ren hong’and’E lian 5’were found.2. DNA was extracted from immature leaves of lotus, and DNA sequences of SSS was amplified by PCR technology. Full-length DNA sequence of SSS was 10 924 bp, consisting of sixteen exons and fifteen introns, among which the third exon （2006 bp） and third intron （1530 bp） were the largest, and while the thirteen exon （99 bp） and second intron （64 bp） are the smallest.3. Expression profiles of SSS gene of lotus were analyzed using the technology of Semi-RT-PCR, and β-actin was used as an internal standard gene. Expression of SSS gene were detected as following tendency in cultivar’ Mei ren hong’:The last leaf> The penultimate leaf > The stalk of the penultimate leaf> rhizome> The stalk of the last leaf. The expression regulation of SSS gene in ’E lian 5’was as follow:The stalk of the last leaf> The last leaf> The penultimate leaf> rhizome> The stalk of the penultimate leaf. The expression of SSS gene in cultivar’Mei ren hong’ was lower than that of cultivar’E lian 5’. The expression of SSS gene was decreased with rhizome growth and developmental process. We found that the mRNA level of SSS gene was 2 times in 1-2 internodes than that of 3-4 internodes. In addition, the expression of SSS gene in cultivar’E lian 5’was 2 times than that of’Mei ren hong’.