The generation of infectious clones is routinely the first step for reverse genetic studies of RNA plant virus gene and sequence function. The procedure given here, details the creation of cDNA clones of tobacco mosaic virus, from which infectious transcripts can be generated in vitro with T7 RNA polymerase. The procedure describes methods for virion purification, viral RNA extraction, reverse transcription, PCR amplification of genomic cDNA fragments, generation of a full-length cDNA clone under the control of a T7 promoter, in vitro transcription, and infectivity testing.
Download full-text PDF |
Source |
|---|---|
| http://dx.doi.org/10.1007/978-1-59745-102-4_32 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Objectives: This study aimed to investigate genotypic characteristics and drug resistance profiles of complex (Mtbc) strains isolated from patients with suspected tuberculosis (TB) in Gabon.
Methods: We performed whole genome sequencing of 430 Mtbc strains cultured between 2012 and 2022. Phylogenetic strain classification, genomic resistance prediction, and cluster analysis were also performed.
Replication-competent recombinant porcine reproductive and respiratory syndrome (PRRS) virus expressing antiviral cytokine interferon-ω5 as a modified live virus vaccine.
Vet Microbiol
January 2025
Department of Agricultural and Environmental Sciences, College of Agriculture, Tennessee State University, Nashville, TN, United States. Electronic address:
Porcine reproductive and respiratory syndrome (PRRS), caused by the highly variable PRRS virus (PRRSV), presents a significant challenge to the swine industry due to its pathogenic and economic burden. The virus evades host immune responses, particularly interferon (IFN) signaling, through various viral mechanisms. Traditional vaccines have shown variable efficacy in the field, prompting the exploration of novel vaccination strategies.
View Article and Find Full Text PDFV6 encoded by mulberry crinkle leaf virus is important for viral DNA replication.
Virology
January 2025
School of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang, Jiangsu, 212100, China; Key Laboratory of Silkworm and Mulberry Genetic Improvement, Ministry of Agriculture and Rural Affairs, Sericultural Scientific Research Center, Chinese Academy of Agricultural Sciences, Zhenjiang, Jiangsu, 212100, China. Electronic address:
Mulberry crinkle leaf virus (MCLV) is a representative species of the genus Mulcrilevirus in the family Geminiviridae. Here, we identified an additional V6 ORF which embedded within the V4 ORF in the MCLV virion-sense strand. The expression of V6 was confirmed by analyzing the promoter activity of V6 ORF upstream sequences and quantifying the viral DNA accumulation in V6-mutant MCLV-infected tomato plants.
View Article and Find Full Text PDFEmerging carbapenem-resistant in a tertiary care hospital in Lima, Peru.
Microbiol Spectr
January 2025
Instituto de Medicina Tropical Alexander von Humboldt, Universidad Peruana Cayetano Heredia, Lima, Peru.
The emergence of carbapenem-resistant (CRKP) poses a significant public health threat, particularly in low- and middle-income countries (LMICs) with limited surveillance and treatment options. This study examines the genetic diversity, resistance patterns, and transmission dynamics of 66 CRKP isolates recovered over 5 years (2015-2019) after the first case of CRKP was identified at a tertiary care hospital in Lima, Peru. Our findings reveal a shift from to as the dominant carbapenemase gene after 2017.
View Article and Find Full Text PDFAn improved reverse genetics system for rotavirus vaccine strain LLR using five plasmid vectors.
J Gen Virol
January 2025
National Key Laboratory of Intelligent Tracking and Forecasting for Infectious Diseases (NITFID), NHC Key Laboratory for Medical Virology and Viral Diseases, National Institute for Viral Disease Control and Prevention, Beijing 100052, PR China.
Species A rotaviruses (RVs), which belong to the family and contain a genome of 11 segmented dsRNA segments, are a leading cause of severe acute gastroenteritis in infants and children younger than 5 years of age. We previously developed a strategy to recover rotavirus vaccine strain LLR from 11 cloned plasmids. Here, we report an improved reverse genetics system for LLR by combining two or three transcriptional cassettes in a single plasmid, which substantially enhances rescue efficiency from 66.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!