Cloning and Expression Analysis of Sugarcane Alkaline/Neutral Invertase Gene SoNIN1

Abstract

Neutral/alkaline invertase irreversibly catalyzes sucrose into glucose and fructose, and plays an important regulating role in plant growth and development. In the present study, the whole length of neutral/alkaline invertase gene cDNA was cloned and sequenced from leaf of sugarcane variety GT28 using RT-PCR(reverse transcription-polymerase chain reaction) and RACE(rapid amplification of cDNA ends), and named as SoNIN1. The gene consists of 2289 bp with an ORF(open reading frame) of 1812 bp encoding a polypeptide of 603 amino acids with calculated protein molecular weight of 67.79 kD, and an isoelectric point of pH 6.41. SoNIN1 gene encodes a protein that is close to that of Zea mays, Oryza sativa, and Lolium perenne, belonging to the same evolutionary branch. The putative protein contains the conserved domain of alkaline/neutral invertase, and no membrane-spanning domain and signal peptide at N-end. The gene promoter sequence was cloned by using genome walking, which contains the basic components of CAAT-boxes and TATA-boxes, as well as specific acting elements such as cis-regulatory element involved in meristem activation, methyl jasmonate response and circadian rhythm, plus drought-induced MYB binding sites, abscisic acid response element, gibberellin response element, etc. Real-time PCR results showed that expression of SoNIN1 could be detected in the roots, stalks, leaves, flowers and buds, among which the highest was found in leaves, the medium in buds, and the lowest in internode +1(enrolled by the top visible dewlap leaf). The expression of SoNIN1 gene in roots and leaves of sugarcane seedling was induced by PEG and NaCl.