Differential Gene Expression Research Of Banana Stressed By Fusarium Oxysporum F.SP.Cubense Using CDNA-AFLP

Fusarium wilt of banana is a destructive vascular systemic fungal disease. In recent years, the disease broke out in the world banana habitat frequently, which had brought the great threat to the banana production and its industry development. Its pathogen is Fusarium oxysporum f. sp. Cubense. The fungus remains in the soil and infects banana plants through the soil, irrigation water and infected suck buds. Now, molecular mechanism of interaction between banana and F. oxysporum has been reported littlely, and the understanding of which would help to research and control the Fusarium wilt of banana. In this study, the interaction mechanism was done by cDNA-AFLP. The results were as following:1,cDNA-AFLP was employed to analyze the differentially expressed genes during compatible interactions between banana and Fusarium wilt. The leaves of banana were inoculated with the adjusted conidial suspension. The controls were inoculated with sterile water. The leaves were collected after4hours,1,3,6days of inoculation and then total RNA was isolated.Double-strained cDNA was synthesized using reverse transcriptase and was digested with EcoR I and Mse I. Specifically designed adaptors were ligated to the ends of restriction fragments. After pre-amplified with primers Epa and Mpa, the templates were diluted20-folds by water and then amplified with selective primers, extended by two selective bases at the3'end.The products were size-fractionated on a6%polyacrylamide sequencing type gel and stained with silver.2,After analysis of cDNA-AFLP,222differentially expressed fragments were obtained. Sequencing and analyzing these fragments,137kinds of positive ESTs were got totally among which64.96%had significant homology with known genes, including12.41%related to primary metabolism,5.84%related to energy metabolism,2.92%related to protein synthesis,5.11%related to protein destination and storage,13.87%related to transcription,4.38%related to transporters,2.19%related to cell growth and division,2.19%related to cell structure,9.48%related to signal transduction,6.57%related to disease and defense response; the remaining35.04% ESTs were unknown about its functions, including24new-found ESTs.3,In this research,9ESTs related to disease and defense response were obtained, including NBS-LRR type resistance gene, hot shock protein, alcohol dehydrogenase and so on;19ESTs related to transcriptional regulation, including the transcription factor MYB and MYC, zinc finger protein, retrotransposon, Glycine-rich RNA binding protein and so on;13ESTs related to signal transduction, including the ubiquitin-conjugating enzyme E2and so on; the remains were related to primary metabolism, energy metabolism and so on.4,The study first successfully applied cDNA-AFLP to analyze the differentially expressed genes during the interactions between banana and F. oxysporum, the partial results of which were verified by semiquantitative RT-PCR. The results of this study will help to reveal the molecular mechanism of banana infected by F. oxysporum, because these genes probably were concerned with the course of the interaction.