Agrobacterium-mediated transformation of grapefruit with the wild-type and mutant RNA-dependent RNA polymerase genes of Citrus tristeza virus Turunçgil tristeza virüsünün (Citrus tristeza virus) normal ve mutant RNA-baǧımlı RNA polimeraz genlerinin Agrobacterium aracılıǧıyla altıntopa aktarılması

ÇEVİK B., Lee R. F., Niblett C. L.

Turkish Journal of Agriculture and Forestry, vol.36, no.2, pp.195-206, 2012 (SCI-Expanded, Scopus, TRDizin) identifier

  • Publication Type: Article / Article
  • Volume: 36 Issue: 2
  • Publication Date: 2012
  • Doi Number: 10.3906/tar-1008-1192
  • Journal Name: Turkish Journal of Agriculture and Forestry
  • Journal Indexes: Science Citation Index Expanded (SCI-EXPANDED), Scopus, TR DİZİN (ULAKBİM)
  • Page Numbers: pp.195-206
  • Keywords: Citrus tristeza virus, Plant transformation, Replicase-mediated resistance, RNA-dependent RNA polymerase
  • Isparta University of Applied Sciences Affiliated: No

Abstract

Citrus paradisi Macfad. 'Duncan' was transformed with constructs coding for the wild-type and mutant RNAdependent RNA polymerase (RdRp) of Citrus tristeza virus (CTV) for exploring replicase-mediated pathogen-derived resistance (RM-PDR). The RdRp gene was amplified from a CTV genome and used to generate the wild-type and 2 mutant RdRp constructs for plant transformation. One mutant had the key amino acids GDD changed to AAA (RdRpmGDD), and the second mutant had a deletion encompassing the GDD domain (RdRp-ΔGDD). Etiolated epicotyl segments of Duncan grapefruit (Citrus paradisi Macfad. 'Duncan') were transformed with each of these constructs using the Agrobacterium-mediated transformation method. From 4540 transformed epicotyl segments, 1402 kanamycinresistant shoots were regenerated. Aft er testing for expression of green fluorescent protein (GFP) and β-glucuronidase (GUS) reporter genes by fluorescence microscopy and histochemical staining, respectively, 146 GUS-positive plants were rooted and 97 surviving plants were established in soil in pots. A total of 70 plants were tested for the presence of the GUS gene and CTV RdRp transgenes by polymerase chain reaction (PCR). A total of 51 GUS and CTV transgenepositive transgenic plants (15 with RdRp, 21 with RdRp-mGDD, and 15 with RdRp-ΔGDD) were identified. © TÜBİTAK.