|Guan, Peizhu - UNIV OF HAWAII|
|Wang, Ming-Li - HARC|
Submitted to: Plant and Animal Genome Conference Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: October 13, 2006
Publication Date: N/A
Technical Abstract: With the genome of papaya (Carica papaya L.) sequenced, the study of gene function is becoming an increasing priority. Our research is to develop an RNA-induced gene silencing tool for the study of functional genomics in papaya. We employed agrobacterium leaf infiltration to induce PTGS in '-glucuronidase (GUS) transgenic (cv Rainbow) and wild type (cv Sunset) papayas. The transient expression of GUS and GFP was achieved by agroinfiltration, indicating agrobacterium infection and gene transfer would occur in infiltrated papaya leaf cells. To determine whether agroinfiltration could be applied to induce PTGS in papaya, GUS and a partial papaya phytoene desaturase (PDS) gene were used as sense, antisense (single-strand) or self-complementary double strand (dsGUS) expression constructs. Agroinfiltration with even empty vector caused loss of GUS staining in the infiltrated zone; for this reason local silencing of GUS could not be detected by histochemical staining. Only one of four rainbow papaya seedlings infiltrated with dsGUS construct showed systemic silencing by histochemical staining; a decrease of GUS mRNA was confirmed by real-time PCR. In papaya infiltrated with PDS expression constructs, no obvious photobleaching occurred locally or systemically. Only one of the plants infiltrated with ssPDS constructs significantly decreased its PDS mRNA level in infiltrated leaves. The results indicate it is possible to use agroinfiltration to induce gene silencing in papaya, but not at a sufficiently efficient or reliable level to be a useful tool for functional genomics. VIGS vectors are currently under investigation as an alternative approach to down-regulation of genes in papaya.