Constructing SSH Library of Cotton Under Drought Stress and Analysis of Drought Associated Genes

Abstract

A forward cDNA-SSH library was constructed from a drought-tolerant cotton (Gossypium hirsutum L.) inbred line, Handan 177, to understand the expressions of drought-induced genes. Using suppression subtractive hybridization method, a total of 2300 positive clones were obtained, of which 300 clones were selected for PCR validation and sequencing. Among these clones, 284 were available sequences. According to clustering analysis of the expressed sequence tag (EST), 202 uniESTs were found, including 28 contigs and 174 singlets. The result of BlastN showed that 156 uniESTs had homologous sequences in GenBank database. The result of BlastX indicated that 116 uniESTs had high homology with proteins with known functions, and 40 uniESTs showed high similarities with unknown proteins or putative proteins. Thirty-three uniESTs were located into 55 KEGG pathways using KOBAS software, including 23 pathways at a significant level (P < 0.05). These significant pathways were mainly related to pyruvate metabolism (15%) and glyoxylate and dicarboxylate metabolism (12%). A large group of drought-induced genes were detected in the cDNA library, which were involved in signal transduction, energy metabolism, protein metabolism, nucleic acid metabolism, photosynthesis, and transmembrane transport. Some of them were associated with drought tolerance, such as malate synthase genes (MS1, 0001_C12 and MS2, 0002_F01), malate dehydrogenase genes (Md1, 001_C12 and Md2, 002_F01), NAC type transcription factor (001_C08), BZR1/BES1 (003_G04), zinc finger protein gene (zfp, 003_C06), and translationally controlled tumor protein gene (tctp, 002_C04).