Identification and analysis of differentially expressed genes during wood formation in Chinese fir by SSH

Abstract

Wood is an important raw material for the global industry with rapidly increasing demand. To isolate the differentially expressed genes in xylogenesis of Chinese fir [Cunninghamia lanceolata (Lamb.) Hook], a forward subtractive cDNA library was constructed using suppression subtractive hybridization (SSH) method, which was performed using the cDNA from the mutant Dugansha clone as the tester and the cDNA from the normal Jurong 0 clone as the driver. Six hundred and eighteen clones were obtained. Recombinants were identified using PCR with universal T7 and SP6 primers and using EcoR digestion. To further eliminate false positive, dot hybridization was used with four DIG-labeled probes (FSP, RSP, UTP, and UDP). Real-time PCR was performed to confirm the results. A total of 260 unique ESTs were obtained, 60% of the ESTs exhibiting homologies with proteins of known function fell into 4 major classes: metabolism, cell wall biogenesis and remodeling, signal transduction and stress. The systematic analysis of genes involved in wood formation in Chinese fir provides valuable insights into the molecular mechanisms involved in xylem differentiation, is important resources for forest research directed toward understanding the genetic control of wood formation and future endeavors to modify wood and fiber properties for industrial use.