Objective: Based on previous study, authors used the suppression subtractive hybridization (SSH) technique to construct the forward and reverse subtractive cDNA libraries of consecutive monoulture Rehmannia glutinosa. Five genes related with consecutive monoculture problem of R. glutinosa were chosen from the each of two subtractive libraries. And their spatiotemporal expression was measured in order to explore the functions in consecutive monoculture problem of R. glutinosa.
Method: Using the real-time quantitative PCR, we tested the relative expression values of the genes in different development stages and tissues of normal growth (one-year culturing) and consecutive monoculture (two-year culturing) R. glutinosa.
Result: The five genes (calcium-dependent protein kinase, s-adenosyl-methionine synthetase, Aminocyclopropane-1-carboxylate oxidase, methyltransferase, calpain), which were chosen from the forward library had high expression in consecutive monoculture R. glutinosa, especially in root, and were hardly expression in normal growth R. glutinosa. On the contrary, the other five genes (RNA-dependent RNA polymerase, RNA replicase, DNA-directed RNA polymerase IIa, cyclin D, RNA binding protein) chosen from the reverse library had high expression in one-year R. glutinosa, but were down regulated or shut down in consecutive monoculture R. glutinosa.
Conclusion: The key genes, which regulate inessential metabolism parthway (such as cyclin D, DNA-directed RNA polymerase IIa), were restrained or shut down in consecutive monoculture R. glutinosa. Calcium and ethylene signaling might played key roles in the formation of consecutive monoculture problem, resulting in disturbing normal metabolic process and syndrome of disease in R. glutinosa appeared in turn.