عنوان

Inhibition of geminiviral replication in a transient assay by expression of tomato leaf curl geminivirusrep gene antisense RNAs

Number

TLpq304393403

.Language of Text, Soundtrack etc

انگلیسی

Title Proper

Inhibition of geminiviral replication in a transient assay by expression of tomato leaf curl geminivirusrep gene antisense RNAs

General Material Designation

[Thesis]

First Statement of Responsibility

O. Chatchawankanphanich

Subsequent Statement of Responsibility

D. P. Maxwell

Name of Publisher, Distributor, etc.

The University of Wisconsin - Madison

Date of Publication, Distribution, etc.

1997

Specific Material Designation and Extent of Item

128

Dissertation or thesis details and type of degree

Ph.D.

Body granting the degree

The University of Wisconsin - Madison

Text preceding or following the note

1997

Text of Note

The genome of tomato leaf curl virus (ToLCV) from Bangalore, India, which is a whitefly-transmitted geminivirus (WTG), was cloned (pIND9) and sequenced. The circular, single-genomic DNA consists of 2,759 nucleotides (GenBank No. U38239), and the genome organization is similar to that of other WTGs with monopartite genomes. It encodes six open reading frames (ORFs), 4 ORFs in the complementary strand and 2 ORFs in the viral strand. Comparison of the nucleotide sequence of pIND9 with those of two other virus isolates from India showed moderate identity 82-87% in the cp (coat protein) gene, but low homology (66-67%) in the origin of replication (ori) and the rep gene (75-81% identity). This virus isolate was not shown to be closely related to other WTGs, by comparison with sequences at the National Center for Biotechnology Information and by phylogenetic trees of the rep and cp genes of 15 WTGs. This virus is considered a distinct monopartite geminivirus and has been given the name ToLCV-Bangalore II. Tomatoes biolistically inoculated with a cloned DNA monomer (pIND9) of ToLCV developed typical leaf curling and yellowing symptoms. The virus was transmitted from biolistically inoculated tomatoes to healthy plants by whiteflies (Bemisia tabaci) and to tomato, Nicotiana benthamiana and Nicotiana tabacum by sap inoculation. Antisense constructs of various regions of the rep gene (c1 ORF) of ToLCV-Bangalore II were cloned into an expression vector. Transient expression in NT-1 tobacco suspension cells has been used to evaluate the efficiency of these antisense constructs for inhibiting ToLCV replication. Tobacco suspension cells were electroporated with the infectious clone of pIND9 and antisense constructs. The antisense construct with the entire rep gene (pAS1-1107) inhibited ToLCV replication by 85% and 100% at the ratio of 1:1 (e.g., 1 mug pIND9-SalI: 1 mug pAS1-1107) and 1:10, respectively. None of the antisense constructs to the partial-length sequence of the rep gene was as effective as pAS1-1107. In addition, pAS1-1107 decreased the replication of other tomato-infecting geminiviruses from Thailand and Egypt at the 1:10 ratio. A duplex of the full-length antisense and sense rep RNAs was detected at 1 day but not 4 days post-inoculation.

Bemisia tabaci

Biological sciences

Nicotiana benthamiana

Nicotiana tabacum

Plant pathology

yellowing

D. P. Maxwell

O. Chatchawankanphanich

Electronic name

p

[Thesis]

276903

a

Y