Obtaining pure cyanophage stocks (plaque purification) v1

protocols.io ◽  
2015 ◽  
Author(s):  
Mathias Middelboe ◽  
Amy M ◽  
and Sif
Keyword(s):  
Plaque Purification

Plaque purification as a method to mitigate the risk of adventitious-agent contamination in influenza vaccine virus seeds

Vaccine ◽  
2011 ◽  
Vol 29 (17) ◽  
pp. 3155-3161 ◽  
Author(s):  
Haruhiko Murata ◽  
Juliete Macauley ◽  
Andrew M. Lewis ◽  
Keith Peden
Keyword(s):  
Influenza Vaccine ◽  
Vaccine Virus ◽  
Plaque Purification

Polymer-Templated Microarrays for Highly Reliable Plaque Purification

2006 ◽  
Vol 22 (1) ◽  
pp. 285-287 ◽  
Author(s):  
K. Na ◽  
M. Lee ◽  
B. Shin ◽  
Y. Je ◽  
J. Hyun
Keyword(s):  
Plaque Purification

scholarly journals High-Frequency Genetic Recombination and Reactivation of Orthopoxviruses from DNA Fragments Transfected into Leporipoxvirus-Infected Cells

2003 ◽  
Vol 77 (13) ◽  
pp. 7281-7290 ◽  
Author(s):  
Xiao-Dan Yao ◽  
David H. Evans
Keyword(s):  
Vaccinia Virus ◽  
Genetic Recombination ◽  
Virus Genome ◽  
Dna Fragments ◽  
Gene Encoding ◽  
Plaque Purification ◽  
Infected Cells ◽  
Clone And Express ◽  
Recombination Reactions ◽  
Multiple Pcr

ABSTRACT Poxvirus DNA is not infectious because establishing an infection requires the activities of enzymes packaged in the virion. This barrier can be overcome by transfecting virus DNA into cells previously infected with another poxvirus, since the resident virus can provide the trans-acting systems needed to reactivate transfected DNA. In this study we show that cells infected with a leporipoxvirus, Shope fibroma virus (SFV), can reactivate vaccinia virus DNA. Similar heterologous packaging systems which used fowlpox-infected cells to reactivate vaccinia virus have been described, but SFV-infected cells promoted a far more efficient reaction that can produce virus titers exceeding 106 PFU/μg of transfected DNA. SFV-promoted reactions also exploit the hyperrecombinogenic systems previously characterized in SFV-infected cells, and these coupled recombination and reactivation reactions could be used to delete nonessential regions of the vaccinia virus genome and to reconstruct vaccinia virus from overlapping DNA fragments. SFV-catalyzed recombination reactions need only two 18- to 20-bp homologies to target PCR amplicons to restriction enzyme-cut vaccinia virus vectors, and this reaction feature was used to rapidly clone and express a gene encoding fluorescent green protein without the need for plaque purification or selectable markers. The ability of SFV-infected cells to reactivate fragments of vaccinia virus was ultimately limited by the number of recombinational exchanges required and one cannot reconstruct vaccinia virus from multiple PCR fragments spanning essential portions of the genome. These observations suggest that recombination is an integral part of poxvirus reactivation reactions and provide a useful new technique for altering the structure of poxvirus genomes.

scholarly journals Virus-induced diabetes mellitus. XVIII. Inhibition by a nondiabetogenic variant of encephalomyocarditis virus.

1980 ◽  
Vol 152 (4) ◽  
pp. 878-892 ◽  
Author(s):  
J W Yoon ◽  
P R McClintock ◽  
T Onodera ◽  
A L Notkins
Keyword(s):  
Infectious Virus ◽  
Islet Cells ◽  
Encephalomyocarditis Virus ◽  
The Other ◽  
Male Mice ◽  
Virus Antibody ◽  
Viral Antigens ◽  
Emc Virus ◽  
Plaque Purification ◽  
Infection Experiments

Plaque purification of the M variant of encephalomyocarditis (EMC) virus resulted in the isolation of two stable variants: one diabetogenic and designated D and the other nondiabetogenic and designated B. When the D variant was inoculated into SJL/J male mice, hypoinsulinemia and hyperglycemia developed in > 90% of the animals. In contrast, none of the mice inoculated with the B variant developed diabetes. Histologic examination of pancreata from mice infected with the D variant revealed insulitis and necrosis of beta cells, whereas islets from mice infected with the B variant showed little, if any, change. When islets were assayed for infectious virus, approximately 10 times more virus was recovered from animals inoculated with the D as compared with the B variant. Moreover, approximately 60% of islet cells from mice infected with the D variant contained viral antigens when stained with fluorescein-labeled anti-EMC virus antibody, whereas < 5% of islet cells from animals infected with the B variant contained viral antigens. Co-infection experiments showed that the induction of diabetes by the D variant was inhibited by the B variant. When the B and D variants were mixed together at B:D ratios of 1, 9, and 99, diabetes developed in 60, 11, and 0% of the mice, respectively. Tissue-culture experiments revealed that the B variant induced considerably more interferon than the D variant, and studies in animals showed that interferon appeared earlier and in greater amounts in the circulation of mice infected with the B as compared with the D variant. These studies suggest that the induction of interferon by the B variant is, at least in part, responsible for the inhibition of diabetes by the D variant.

scholarly journals Effect of the Selection Pressure of Vaccine Antibodies on Evolution of H9N2 Avian Influenza Virus in Chickens

2020 ◽  
Author(s):  
Hailong SU ◽  
Yu Zhao ◽  
Lirong Zheng ◽  
Shifeng Wang ◽  
Huoying Shi ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
High Frequency ◽  
Selection Pressure ◽  
H9n2 Avian Influenza Virus ◽  
Gene Variation ◽  
Fifth Generation ◽  
Plaque Purification ◽  
Prevention Method

Abstract H9N2 avian influenza virus has spread worldwide, and vaccination with an inactivated virus is currently the major prevention method in China. To further understand the effect of the selection pressure from antibodies on the evolution of H9N2 avian influenza virus, F/98 (A/Chicken/Shanghai/F/98), which is the vaccine representative of H9N2 avian influenza virus in East China, was used for serial passaging for 20 generations in chickens with and without vaccination. After plaque purification from trachea and lung tissues, 390 quasispecies were obtained. The second-generation quasispecies under the selection pressure of vaccine antibodies had undergone 100% antigen variation, while after passaging to the fifth generation, only 30-40% of the quasispecies displayed antigen variation when there was no selection pressure of vaccine antibodies, implying that the selection pressure of vaccine antibodies promotes the antigen variation of F/98. We found for the first time that there were three mutation hotspots in the HA genes of the quasispecies under the selection pressure of vaccine antibodies, which were K131R, A168T, and N201D. Moreover, under the selection pressure of vaccine antibodies, 10 amino acids (67-76) of the NA protein of all quasispecies were deleted, and PB2 of the quasispecies had undergone a high-frequency R355K mutation. However, without selection pressure of vaccine antibodies, NP had undergone two high-frequency mutations, namely, V186I and L466I, and a high-frequency mutation of L77I appeared in the NS gene. This result shows that the vaccine antibody selection pressure could control and regulate gene variation of the F/98 virus. Compared to that of the parental virus F/98, the EID 50 of the twentieth passaged virus under the selection pressure of vaccine antibodies did not change, while the EID 50 of the twentieth passaged virus without selection pressure of vaccine antibodies was significantly enhanced by 794 times. Furthermore, the twentieth passaged virus with selection pressure from vaccine antibodies lost its lethal ability in embryonated chicken eggs, whereas the EID 50 of the twentieth passaged virus without selection pressure of vaccine antibodies increased to 6.3 times that of the F/98 strain. All the above results show that the selection pressure of vaccine antibodies promotes the antigen variation of H9N2 avian influenza virus and plays a role in regulating and controlling gene mutation of H9N2 avian influenza virus.

scholarly journals Plaque Purification of Bluetongue Virus Serotype-4 (BTV-4)

2018 ◽  
Vol 7 (04) ◽  
pp. 2837-2844
Author(s):  
M. Srikanth Reddy ◽  
Kalyani Putty ◽  
Y. Narasimha Reddy ◽  
P.P. Rao ◽  
M. Ramakoti Reddy ◽  
...  
Keyword(s):  
Bluetongue Virus ◽  
Virus Serotype ◽  
Plaque Purification

scholarly journals Effect of the Selection Pressure of Vaccine Antibodies on Evolution of H9N2 Avian Influenza Virus in Chickens

2020 ◽  
Author(s):  
Hailong SU ◽  
Yu Zhao ◽  
Lirong Zheng ◽  
Shifeng Wang ◽  
Huoying Shi ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
High Frequency ◽  
Selection Pressure ◽  
H9n2 Avian Influenza Virus ◽  
Gene Variation ◽  
Fifth Generation ◽  
Plaque Purification ◽  
Prevention Method

Abstract H9N2 avian influenza virus has spread worldwide, and vaccination with an inactivated virus is currently the major prevention method in China. To further understand the effect of the selection pressure from antibodies on the evolution of H9N2 avian influenza virus, F/98 (A/Chicken/Shanghai/F/98), which is the vaccine representative of H9N2 avian influenza virus in East China, was used for serial passaging for 20 generations in chickens with and without vaccination. After plaque purification from trachea and lung tissues, 390 quasispecies were obtained. The second-generation quasispecies under the selection pressure of vaccine antibodies had undergone 100% antigen variation, while after passaging to the fifth generation, only 30-40% of the quasispecies displayed antigen variation when there was no selection pressure of vaccine antibodies, implying that the selection pressure of vaccine antibodies promotes the antigen variation of F/98. We found for the first time that there were three mutation hotspots in the HA genes of the quasispecies under the selection pressure of vaccine antibodies, which were K131R, A168T, and N201D. Moreover, under the selection pressure of vaccine antibodies, 10 amino acids (67-76) of the NA protein of all quasispecies were deleted, and PB2 of the quasispecies had undergone a high-frequency R355K mutation. However, without selection pressure of vaccine antibodies, NP had undergone two high-frequency mutations, namely, V186I and L466I, and a high-frequency mutation of L77I appeared in the NS gene. This result shows that the vaccine antibody selection pressure could control and regulate gene variation of the F/98 virus. Compared to that of the parental virus F/98, the EID 50 of the twentieth passaged virus under the selection pressure of vaccine antibodies did not change, while the EID 50 of the twentieth passaged virus without selection pressure of vaccine antibodies was significantly enhanced by 794 times. Furthermore, the twentieth passaged virus with selection pressure from vaccine antibodies lost its lethal ability in embryonated chicken eggs, whereas the EID 50 of the twentieth passaged virus without selection pressure of vaccine antibodies increased to 6.3 times that of the F/98 strain. All the above results show that the selection pressure of vaccine antibodies promotes the antigen variation of H9N2 avian influenza virus and plays a role in regulating and controlling gene mutation of H9N2 avian influenza virus.

scholarly journals Hobi-like pestivirus: both biotypes isolated from a diseased animal

2012 ◽  
Vol 93 (9) ◽  
pp. 1976-1983 ◽  
Author(s):  
Nicola Decaro ◽  
Viviana Mari ◽  
Pierfrancesco Pinto ◽  
Maria Stella Lucente ◽  
Rossana Sciarretta ◽  
...  
Keyword(s):  
Molecular Characterization ◽  
Respiratory Disease ◽  
Full Length ◽  
Kidney Cells ◽  
Diseased Animal ◽  
Bovine Kidney ◽  
End Point ◽  
Plaque Purification

A Hobi-like pestivirus pair consisting of cytopathogenic (cp) and non-cytopathogenic (noncp) strains, Italy 83/10cp and Italy 83/10ncp, was isolated from the lung of a heifer that died of respiratory disease. The noncp and cp viruses were isolated on Madin–Darby bovine kidney cells and separated by plaque purification and end point dilution. Analysis of the nearly full-length genomes revealed that the two viruses were very closely related to each other and to the noncp Hobi-like strain Italy 1/10-1, which had been isolated a few weeks earlier from the same herd. One major difference between noncp and cp viruses concerned the presence of a cellular Jiv sequence in the 3′ domain of the NS2-encoding region of the cp strain. This is the first study, to our knowledge, reporting the isolation and molecular characterization of a Hobi-like virus pair.

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