Construction and Application of a Baculovirus Genomic Library

Chen, Yin; Lin, Xiaojie; Yao, Yi; Lu, Yiyu; Zhang, Zhifang

doi:10.1515/znc-2009-7-817

Cited by 9 publications

(6 citation statements)

References 18 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The precipitate was washed several times with distilled water and re-suspended in 0.1% SDS for 30 min at room temperature. After centrifugation, the clean polyhedra were re-suspended in 200 μl TE buffer (10 mM Tris–HCl, pH 8.0, 1 mM EDTA) [15]. …”

Section: Methodsmentioning

confidence: 99%

“…A random genomic library of HearMNPV was constructed according to the “partial filling-in” method and contained 2.0 to 5.0 kbp fragment in vector pUC19 [15,16] DNA fragments for sequencing were prepared from 527 recombinant plasmids. The recombinant plasmids were sequenced with plasmid specific primers and 'primer nesting' from both strands, using BigDye Terminator v3.1 (ABI) on a 3130XL Genetic analyzer (ABI).…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Genomic sequencing and analyses of HearMNPV—a new Multinucleocapsid nucleopolyhedrovirus isolated from Helicoverpa armigera

Tang

Zhang

et al. 2012

Virol J

Self Cite

View full text Add to dashboard Cite

BackgroundHearMNPV, a nucleopolyhedrovirus (NPV), which infects the cotton bollworm, Helicoverpa armigera, comprises multiple rod-shaped nucleocapsids in virion(as detected by electron microscopy). HearMNPV shows a different host range compared with H. armigera single-nucleocapsid NPV (HearSNPV). To better understand HearMNPV, the HearMNPV genome was sequenced and analyzed.MethodsThe morphology of HearMNPV was observed by electron microscope. The qPCR was used to determine the replication kinetics of HearMNPV infectious for H. armigera in vivo. A random genomic library of HearMNPV was constructed according to the “partial filling-in” method, the sequence and organization of the HearMNPV genome was analyzed and compared with sequence data from other baculoviruses.ResultsReal time qPCR showed that HearMNPV DNA replication included a decreasing phase, latent phase, exponential phase, and a stationary phase during infection of H. armigera. The HearMNPV genome consists of 154,196 base pairs, with a G + C content of 40.07%. 162 putative ORFs were detected in the HearMNPV genome, which represented 90.16% of the genome. The remaining 9.84% constitute four homologous regions and other non-coding regions. The gene content and gene arrangement in HearMNPV were most similar to those of Mamestra configurata NPV-B (MacoNPV-B), but was different to HearSNPV. Comparison of the genome of HearMNPV and MacoNPV-B suggested that HearMNPV has a deletion of a 5.4-kb fragment containing five ORFs. In addition, HearMNPV orf66, bro genes, and hrs are different to the corresponding parts of the MacoNPV-B genome.ConclusionsHearMNPV can replicate in vivo in H. armigera and in vitro, and is a new NPV isolate distinguished from HearSNPV. HearMNPV is most closely related to MacoNPV-B, but has a distinct genomic structure, content, and organization.

show abstract

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Genomic sequencing and analyses of HearMNPV—a new Multinucleocapsid nucleopolyhedrovirus isolated from Helicoverpa armigera

Tang

Zhang

et al. 2012

Virol J

Self Cite

View full text Add to dashboard Cite

show abstract

“…A random genomic library of SpltNPV-C3 was constructed according to the partial filling-in method (Chen et al, 2009) [ 9 ]. ORFs were defined using ORF Finder ( http://www.ncbi.nlm.nih.gov/gorf/gorf.html ).…”

Section: Methodsmentioning

confidence: 99%

Genome characteristics and the ODV proteome of a second distinct alphabaculovirus from Spodoptera litura

Gao,

Liu,

Gao

et al. 2024

BMC Genomics

Self Cite

View full text Add to dashboard Cite

Background Spodoptera litura is a harmful pest that feeds on more than 80 species of plants, and can be infected and killed by Spodoptera litura nucleopolyhedrovirus (SpltNPV). SpltNPV-C3 is a type C SpltNPV clone, that was observed and collected in Japan. Compared with type A or type B SpltNPVs, SpltNPV-C3 can cause the rapid mortality of S. litura larvae. Methods In this study, occlusion bodies (OBs) and occlusion-derived viruses (ODVs) of SpltNPV-C3 were purified, and OBs were observed by scanning electron microscopy (SEM). ODVs were observed under a transmission electron microscope (TEM). Results Both OBs and ODVs exhibit morphological characteristics typical of nucleopolyhedroviruses (NPVs).The genome of SpltNPV-C3 was sequenced and analyzed; the total length was 148,634 bp (GenBank accession 780,426,which was submitted as SpltNPV-II), with a G + C content of 45%. A total of 149 predicted ORFs were found. A phylogenetic tree of 90 baculoviruses was constructed based on core baculovirus genes. LC‒MS/MS was used to analyze the proteins of SpltNPV-C3; 34 proteins were found in the purified ODVs, 15 of which were core proteins. The structure of the complexes formed by per os infectivity factors 1, 2, 3 and 4 (PIF-1, PIF-2, PIF-3 and PIF-4) was predicted with the help of the AlphaFold multimer tool and predicted conserved sequences in PIF-3. SpltNPV-C3 is a valuable species because of its virulence, and the analysis of its genome and proteins in this research will be beneficial for pest control efforts.

show abstract

“…The details for cell culture were from Summers and Smith's manual [ 15 ]. A random genomic library of BmNPV was constructed according to the "partial filling-in" method that contained a 3 kb to 5 kb fragment in the pUC19 vector [ 16 , 17 ]. Plasmid DNAs of 238 positive colonies were extracted for further transient assays [ 17 ].…”

Section: Methodsmentioning

confidence: 99%

Baculovirus immediately early 1, a mediator for homologous regions enhancer function in trans

Lin

Chen

Yao

et al. 2010

Virol J

Self Cite

View full text Add to dashboard Cite

BackgroundEnhancers are DNA sequences that serve as binding sites for regulatory proteins, and stimulate transcriptional activity independent of their positions and orientations with respect to the transcriptional initiation site. Previous studies considered that baculovirus homologous regions (hrs) function as enhancers in cis. In our study, a plasmid containing homologous region 3 (hr3) enhancer from Bombyx mori nucleopolyhedrovirus (BmNPV) failed to enhance transcription of promoter in other plasmid in co-transfection assays, but strong stimulation occurred when cells were infected by BmNPV. ResultsThe cotransfection results of each BmNPV genomic library plasmid, hr3 plasmid and reporter plasmid showed that there were eight library plasmids stimulated the luciferase gene expression remarkably. Sequencing these plasmids revealed that each of them contained the ie-1 gene. Transfected plasmids, containing ie-1, hr3 and various origin promoter drove reporter gene showed the function was even retained. Cotransfection of hr3 functional dissected fragment and ie-1 revealed that the 30-bp imperfect palindrome destroyed fragment can't enhance reporter gene expression even though transfected with ie-1.ConclusionIE-1 was the only early factor of BmNPV that could act as a mediator for hr enhancer function in trans and the trans-function was achieved with a broad-spectrum of promoters. The 30-bp imperfect palindrome was the elementary molecular structure by which IE-1 participated in the enhancer function in trans.

show abstract

Construction and Application of a Baculovirus Genomic Library

Cited by 9 publications

References 18 publications

Genomic sequencing and analyses of HearMNPV—a new Multinucleocapsid nucleopolyhedrovirus isolated from Helicoverpa armigera

Genomic sequencing and analyses of HearMNPV—a new Multinucleocapsid nucleopolyhedrovirus isolated from Helicoverpa armigera

Genome characteristics and the ODV proteome of a second distinct alphabaculovirus from Spodoptera litura

Baculovirus immediately early 1, a mediator for homologous regions enhancer function in trans

Contact Info

Product

Resources

About