scholarly journals Large-scale RT-PCR recovery of full-length cDNA clones

BioTechniques ◽  
2004 ◽  
Vol 36 (4) ◽  
pp. 690-700 ◽  
Author(s):  
Jia Qian Wu ◽  
Angela M. Garcia ◽  
Steven Hulyk ◽  
Anna Sneed ◽  
Carla Kowis ◽  
...  
Keyword(s):  
Large Scale ◽  
Full Length ◽  
Cdna Clones ◽  
Rt Pcr ◽  
Full Length Cdna

Cloning and characterization of full-length cDNA clones encoding chalcone synthase from the orchid Bromheadia finlaysoniana

1998 ◽  
Vol 36 (9) ◽  
pp. 647-656 ◽  
Author(s):  
Chye-Fong Liew ◽  
Chong-Jin Goh ◽  
Chiang-Shiong Loh ◽  
Saw-Hoon Lim
Keyword(s):  
Chalcone Synthase ◽  
Full Length ◽  
Cdna Clones ◽  
Full Length Cdna

scholarly journals Large-scale collection of full-length cDNA and transcriptome analysis inHevea brasiliensis

DNA Research ◽  
2017 ◽  
pp. dsw056 ◽  
Author(s):  
Yuko Makita ◽  
Kiaw Kiaw Ng ◽  
G. Veera Singham ◽  
Mika Kawashima ◽  
Hideki Hirakawa ◽  
...  
Keyword(s):  
Transcriptome Analysis ◽  
Large Scale ◽  
Full Length ◽  
Full Length Cdna

scholarly journals Infectious Transcripts from Cloned Genome-Length cDNA of Porcine Reproductive and Respiratory Syndrome Virus

1998 ◽  
Vol 72 (1) ◽  
pp. 380-387 ◽  
Author(s):  
J. J. M. Meulenberg ◽  
J. N. A. Bos-de Ruijter ◽  
R. van de Graaf ◽  
G. Wensvoort ◽  
R. J. M. Moormann
Keyword(s):  
Cdna Clone ◽  
De Novo ◽  
Infectious Clone ◽  
Full Length ◽  
Respiratory Syndrome Virus ◽  
Cdna Clones ◽  
Genome Length ◽  
Full Length Cdna ◽  
Lung Macrophages ◽  
Growth Properties

ABSTRACT The 5′-terminal end of the genomic RNA of the Lelystad virus isolate (LV) of porcine reproductive and respiratory syndrome virus was determined. To construct full-length cDNA clones, the 5′-terminal sequence was ligated to cDNA clones covering the complete genome of LV. When RNA that was transcribed in vitro from these full-length cDNA clones was transfected into BHK-21 cells, infectious LV was produced and secreted. The virus was rescued by passage to porcine alveolar lung macrophages or CL2621 cells. When infectious transcripts were transfected to porcine alveolar lung macrophages or CL2621 cells, no infectious virus was produced due to the poor transfection efficiency of these cells. The growth properties of the viruses produced by BHK-21 cells transfected with infectious transcripts of LV cDNA resembled the growth properties of the parental virus from which the cDNA was derived. Two nucleotide changes leading to a unique PacI restriction site directly downstream of the ORF7 gene were introduced in the genome-length cDNA clone. The virus recovered from this mutated cDNA clone retained the PacI site, which confirmed the de novo generation of infectious LV from cloned cDNA. These results indicate that the infectious clone of LV enables us to mutagenize the viral genome at specific sites and that it will therefore be useful for detailed molecular characterization of the virus, as well as for the development of a safe and effective live vaccine for use in pigs.

Production of full-length cDNA from a picornaviral genome by RT-PCR

1996 ◽  
Vol 12 (1) ◽  
pp. 11 ◽  
Author(s):  
Dario Leister ◽  
Richard Thompson ◽  
Otfried Marquardt
Keyword(s):  
Full Length ◽  
Rt Pcr ◽  
Full Length Cdna

scholarly journals A Reverse Genetics System for Zika Virus Based on a Simple Molecular Cloning Strategy

Viruses ◽  
2018 ◽  
Vol 10 (7) ◽  
pp. 368 ◽  
Author(s):  
Maximilian Münster ◽  
Anna Płaszczyca ◽  
Mirko Cortese ◽  
Christopher Neufeldt ◽  
Sarah Goellner ◽  
...  
Keyword(s):  
Zika Virus ◽  
In Silico ◽  
Reverse Genetics ◽  
Molecular Mechanisms ◽  
Full Length ◽  
Cdna Clones ◽  
In Silico Prediction ◽  
Fluorescent Reporter ◽  
Full Length Cdna ◽  
Spurious Transcription

The Zika virus (ZIKV) has recently attracted major research interest as infection was unexpectedly associated with neurological manifestations in developing foetuses and with Guillain-Barré syndrome in infected adults. Understanding the underlying molecular mechanisms requires reverse genetic systems, which allow manipulation of infectious cDNA clones at will. In the case of flaviviruses, to which ZIKV belongs, several reports have indicated that the construction of full-length cDNA clones is difficult due to toxicity during plasmid amplification in Escherichia coli. Toxicity of flaviviral cDNAs has been linked to the activity of cryptic prokaryotic promoters within the region encoding the structural proteins leading to spurious transcription and expression of toxic viral proteins. Here, we employ an approach based on in silico prediction and mutational silencing of putative promoters to generate full-length cDNA clones of the historical MR766 strain and the contemporary French Polynesian strain H/PF/2013 of ZIKV. While for both strains construction of full-length cDNA clones has failed in the past, we show that our approach generates cDNA clones that are stable on single bacterial plasmids and give rise to infectious viruses with properties similar to those generated by other more complex assembly strategies. Further, we generate luciferase and fluorescent reporter viruses as well as sub-genomic replicons that are fully functional and suitable for various research and drug screening applications. Taken together, this study confirms that in silico prediction and silencing of cryptic prokaryotic promoters is an efficient strategy to generate full-length cDNA clones of flaviviruses and reports novel tools that will facilitate research on ZIKV biology and development of antiviral strategies.

scholarly journals Genetic Manipulation of Equine Arteritis Virus Using Full-Length cDNA Clones: Separation of Overlapping Genes and Expression of a Foreign Epitope

Virology ◽  
2000 ◽  
Vol 270 (1) ◽  
pp. 84-97 ◽  
Author(s):  
Antoine A.F. de Vries ◽  
Amy L. Glaser ◽  
Martin J.B. Raamsman ◽  
Cornelis A.M. de Haan ◽  
Sonia Sarnataro ◽  
...  
Keyword(s):  
Genetic Manipulation ◽  
Full Length ◽  
Equine Arteritis Virus ◽  
Cdna Clones ◽  
Overlapping Genes ◽  
Full Length Cdna ◽  
Foreign Epitope

scholarly journals RNA transcripts of full-length cDNA clones of rabbit hepatitis E virus are infectious in rabbits

2015 ◽  
Vol 96 (5) ◽  
pp. 1190-1190
Author(s):  
Caitlin M. Cossaboom ◽  
Yao-Wei Huang ◽  
Danielle M. Yugo ◽  
Scott P. Kenney ◽  
Pablo Piñeyro ◽  
...  
Keyword(s):  
Hepatitis E Virus ◽  
Hepatitis E ◽  
Full Length ◽  
Cdna Clones ◽  
Full Length Cdna ◽  
Rna Transcripts
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