Molecular cloning and comparative sequence analyses of bluetongue virus S1 segments by selective synthesis of specific full-length DNA copies of dsRNA genes

Timothy F. Kowalik, Yi Yuan Yang, Joseph K K Li

研究成果: 雜誌貢獻文章

43 引文 (Scopus)

摘要

Using primers complementary to the conserved sequences of the 3′ ends of bluetongue virus genomic dsRNA segments, full-length DNA clones of all 10 dsRNA genes from the five U.S. BTV serotypes were synthesized and amplified by a novel method (CIampR). This amounts to nearly 100,000 base pairs of dsRNA cloned as unique full-length DNA copies. This continuous one-tube procedure combined cloning of the denatured dsRNA with reverse transcriptase and the selective amplification of full-length DNA by the polymerase chain reaction. CIampR-derived clones of the genomic segment S1 of BTV-11 encoding the serogroup antigen, VP7, were sequenced and shown to be complete copy, containing a total of 1156 by and a long open reading frame of 349 amino acids. Comparative sequence analyses of BTV-11 S1 with those of the other U.S. serotypes show that 95.2% of the nucleotides are conserved between BTV-11 and -10, while only 79.0% of the bases are identical between BTV-11 and -13. Comparison of the VP7 proteins demonstrates that 100% of the amino acids are conserved between BTV-11 and -10 and 93.7% of these residues are identical between VP7 of BTV-11 and -13. The adaptation of the polymerase chain reaction to the full-length cloning and amplification of dsRNA (CIampR) should greatly facilitate molecular studies within the Reoviridae family.

原文英語
頁(從 - 到)820-823
頁數4
期刊Virology
177
發行號2
DOIs
出版狀態已發佈 - 1990
對外發佈Yes

指紋

Bluetongue virus
Molecular Cloning
Sequence Analysis
Organism Cloning
DNA
Genes
Clone Cells
Reoviridae
Amino Acids
Polymerase Chain Reaction
Conserved Sequence
RNA-Directed DNA Polymerase
DNA-Directed DNA Polymerase
Base Pairing
Open Reading Frames
Nucleotides
Antigens
Serogroup
Proteins

ASJC Scopus subject areas

  • Virology
  • Infectious Diseases

引用此文

Molecular cloning and comparative sequence analyses of bluetongue virus S1 segments by selective synthesis of specific full-length DNA copies of dsRNA genes. / Kowalik, Timothy F.; Yang, Yi Yuan; Li, Joseph K K.

於: Virology, 卷 177, 編號 2, 1990, p. 820-823.

研究成果: 雜誌貢獻文章

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abstract = "Using primers complementary to the conserved sequences of the 3′ ends of bluetongue virus genomic dsRNA segments, full-length DNA clones of all 10 dsRNA genes from the five U.S. BTV serotypes were synthesized and amplified by a novel method (CIampR). This amounts to nearly 100,000 base pairs of dsRNA cloned as unique full-length DNA copies. This continuous one-tube procedure combined cloning of the denatured dsRNA with reverse transcriptase and the selective amplification of full-length DNA by the polymerase chain reaction. CIampR-derived clones of the genomic segment S1 of BTV-11 encoding the serogroup antigen, VP7, were sequenced and shown to be complete copy, containing a total of 1156 by and a long open reading frame of 349 amino acids. Comparative sequence analyses of BTV-11 S1 with those of the other U.S. serotypes show that 95.2{\%} of the nucleotides are conserved between BTV-11 and -10, while only 79.0{\%} of the bases are identical between BTV-11 and -13. Comparison of the VP7 proteins demonstrates that 100{\%} of the amino acids are conserved between BTV-11 and -10 and 93.7{\%} of these residues are identical between VP7 of BTV-11 and -13. The adaptation of the polymerase chain reaction to the full-length cloning and amplification of dsRNA (CIampR) should greatly facilitate molecular studies within the Reoviridae family.",
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