Genetic Transformation of Citrus paradisi with Antisense and Untranslatable RNA-dependent RNA Polymerase Genes of Citrus tristeza closterovirus

Bayram ÇEVİK; Richard F. LEE; Charles L. NIBLETT

Makale detayları

Genetic Transformation of Citrus paradisi with Antisense and Untranslatable RNA-dependent RNA Polymerase Genes of Citrus tristeza closterovirus

Yazarlar:

Bayram ÇEVİK | Richard F. LEE | Charles L. NIBLETT

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Agriculture and Forestry 2006 Vol.30 Issue 3, pp.173-182

Belge Tipi:

Makale

Yayın Tarihi:

2/2006

Belge Dili:

Türkçe

Document ID (Org No.):

2000808880

Sıklık:

İki Aylık

eISSN:

1303-6173

pISSN:

1300-011X

Anahtar kelimeler:

Özet:

 

Protein and RNA-mediated forms of pathogen-derived resistance (PDR) have been developed against many viruses in different plants. However, no resistance has been reported against Citrus tristeza virus (CTV), a closterovirus, in Citrus species transformed with coat protein genes or other sequences of CTV. The successful use of replication-associated genes in RNA-mediated resistance in other crops prompted the use of the RNA-dependent RNA polymerase (RdRp) gene of CTV for the development of RNA-mediated PDR in Citrus. The RdRP gene was amplified from CTV isolate DPI3800 from Florida and used to generate antisense (RdRp-AS) and untranslatable (RdRp-UT) constructs with point mutation consecutive stop codons in the 5’ end of the RdRp gene for use in plant transformation. A total of 3120 etiolated epicotyl segments of Duncan grapefruit (Citrus paradisi Macf. cv. Duncan) were transformed with these constructs using Agrobacterium tumefaciens-mediated transformation. From these segments 1040 kanamycin-resistant shoots were regenerated, and a total of 131 putative transgenic shoots were identified by fluorescent microscopy and histochemical b-glucuronidase (GUS) assays. One hundred GUS positive plants were rooted and 66 plants survived and were established on soil. A total of 41 plants were tested by polymerase chain reaction (PCR) for the presence of the GUS gene and for the transgenes. Eighteen GUS-positive and transgene-positive plants (8 with RdRp-AS, and 10 with RdRp-UT) were identified.

 

Protein and RNA-mediated forms of pathogen-derived resistance (PDR) have been developed against many viruses in different plants. However, no resistance has been reported against Citrus tristeza virus (CTV), a closterovirus, in Citrus species transformed with coat protein genes or other sequences of CTV. The successful use of replication-associated genes in RNA-mediated resistance in other crops prompted the use of the RNA-dependent RNA polymerase (RdRp) gene of CTV for the development of RNA-mediated PDR in Citrus. The RdRP gene was amplified from CTV isolate DPI3800 from Florida and used to generate antisense (RdRp-AS) and untranslatable (RdRp-UT) constructs with point mutation consecutive stop codons in the 5’ end of the RdRp gene for use in plant transformation. A total of 3120 etiolated epicotyl segments of Duncan grapefruit (Citrus paradisi Macf. cv. Duncan) were transformed with these constructs using Agrobacterium tumefaciens-mediated transformation. From these segments 1040 kanamycin-resistant shoots were regenerated, and a total of 131 putative transgenic shoots were identified by fluorescent microscopy and histochemical b-glucuronidase (GUS) assays. One hundred GUS positive plants were rooted and 66 plants survived and were established on soil. A total of 41 plants were tested by polymerase chain reaction (PCR) for the presence of the GUS gene and for the transgenes. Eighteen GUS-positive and transgene-positive plants (8 with RdRp-AS, and 10 with RdRp-UT) were identified.

Makale detayları

Genetic Transformation of Citrus paradisi with Antisense and Untranslatable RNA-dependent RNA Polymerase Genes of Citrus tristeza closterovirus

Yazarlar:

Bayram ÇEVİK | Richard F. LEE | Charles L. NIBLETT

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Agriculture and Forestry 2006 Vol.30 Issue 3, pp.173-182

Belge Tipi:

Makale

Yayın Tarihi:

2/2006

Belge Dili:

Türkçe

Document ID (Org No.):

2000808880

Sıklık:

İki Aylık

eISSN:

1303-6173

pISSN:

1300-011X

Citrus tristeza virus

replicase and RNA-mediated resistance

plant transformation

RNA-dependent RNA polymerase

Sayı

Makale detayları

Genetic Transformation of Citrus paradisi with Antisense and Untranslatable RNA-dependent RNA Polymerase Genes of Citrus tristeza closterovirus

Yazarlar: Bayram ÇEVİK | Richard F. LEE | Charles L. NIBLETT

Yayıncı: TUBITAK

Dergi: Turkish Journal of Agriculture and Forestry 2006 Vol.30 Issue 3, pp.173-182

Belge Tipi: Makale

Yayın Tarihi: 2/2006

Belge Dili: Türkçe

Document ID (Org No.): 2000808880

Sıklık: İki Aylık

eISSN: 1303-6173

pISSN: 1300-011X

Citrus tristeza virus

replicase and RNA-mediated resistance

plant transformation

RNA-dependent RNA polymerase

Sayı

Yazarlar:

Bayram ÇEVİK | Richard F. LEE | Charles L. NIBLETT

Yayıncı:

TUBITAK

Dergi:

Turkish Journal of Agriculture and Forestry 2006 Vol.30 Issue 3, pp.173-182

Belge Tipi:

Makale

Yayın Tarihi:

2/2006

Belge Dili:

Türkçe

Document ID (Org No.):

2000808880

Sıklık:

İki Aylık

eISSN:

1303-6173

pISSN:

1300-011X