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Title: RNA-INDUCED GENE SILENCING IN PAPAYAS

Author
item GUAN, P - UNIV HAWAII
item Moore, Paul
item Albert, Henrik

Submitted to: CTAHR Student Research Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 3/3/2006
Publication Date: 4/7/2006
Citation: Guan, P., Moore, P.H., Albert, H.H. 2006. RNA-induced gene silencing in papayas. 18th Annual CTAHR Student Research Symposium. (Abstract)#51. Pg. 63.

Interpretive Summary:

Technical Abstract: (ab) Agrobacterium leaf infiltration is a widely used method for inducing post-transcriptional gene silencing (PTGS) in Nicotiana benthamiana but has rarely been applied successfully in other species. Here we employed agrobacterium leaf infiltration to induce PTGS in ß-glucuronidase (GUS) transgenic (cv SunUp, cv Rainbow) and wild type (cv Sunset) papayas. To determine whether agrobacterium infection and gene transfer would occur in infiltrated papaya leaf cells, GUS and GFP reporter constructs were tested. To determine whether leaf infiltration could be applied to induce PTGS in papayas, GUS and a partial papaya phytoene desaturase (PDS) gene, were used as sense, antisense (single-strand) or self-complementary double strand (dsGUS) expression constructs. In SunUp (homozygous) papayas infiltrated with dsGUS, histochemical staining became faint on infiltrated leaves at 4 days post-infiltration (dpi), indicating GUS activity was reduced. Later, newly emerged leaves showed a significant decline of GUS expression as compared to controls. In Rainbow (heterozygous) papayas, no local silencing was found, but one,among four rainbow papaya seedlings infiltrated with dsGUS construct, showed systemic silencing at 9 dpi; quantitative RT-PCR indicated GUS mRNA levels were decreased by approximately 50%. In papayas infiltrated with PDS expression constructs, sense,antisense, or double-stranded, no obvious photobleaching occurred locally or systemically. Only one of the plants infiltrated with PDS 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 sufficiently efficient or reliable to be a useful tool for functional genomics. Development of an alternative method for down-regulating genes in papaya will be an essential step in utilizing the whole genome sequence data currently being generated for this crop.