Efficient bacterial expression of recombinant potato mop-top virus non-structural triple gene block protein 1 modified by progressive deletion of its N-terminus
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61389030%3A_____%2F05%3A00028432" target="_blank" >RIV/61389030:_____/05:00028432 - isvavai.cz</a>
Result on the web
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DOI - Digital Object Identifier
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Alternative languages
Result language
angličtina
Original language name
Efficient bacterial expression of recombinant potato mop-top virus non-structural triple gene block protein 1 modified by progressive deletion of its N-terminus
Original language description
To obtain strong bacterial expression of proteins that seem to be hard to express in bacteria or are highly toxic for bacteria, it is possible to vreate a palette of similar constructs, differing only by several nucleotides, gradually deleted from the full-lenght clone by ewonuclease III. When a construct is equipped with the 6xHis tag, a simple colony-blot procedure can be performed and a colony giving strong and efficient expression can easily sbe selected for high range protein expression. We utilized this procedure to produce one of potato mop-top virus (PMTV) movement proteins, namely triple gene block protein 1 (TGPp1) which was very hard toexpress in bacteria in its original length. The TGPp1 gene was digested with exonuclease III and nuclease S1 from its 5´ terminus. Leaving 6xHis tag intact. The clone that showed the strongest signal with anti-His antibodies in colony-blot procedure was found to have 44 amino acids (of total 463) deleted. The SDS-PAGE and Western blot of h...
Czech name
Účinná bakteriální exprese rekombinantního nestrukturálního proteinu 1 z ?triple gene blocku? mop-top viru bramboru
Czech description
Byla vypracována metoda pro expresi těžko exprimovatelných či toxických proteinů v bakteriích. Pomocí postupného zkracování exonukleasou III a nukleasou S1 z 5´konce se vytvořila škála podobných konstruktů lišících se pouze o několik nukleotidů. Konstrukty byly vybaveny 6 x His kotvou, která umožnila jejich detekci v colony- blotu. Pro expresi ve velkém měřítku se poté vybraly kolonie poskytující dostatečně silnou expresi. Tato metoda byla využita pro přípravu jednoho z nestrukturních ?movement? proteinů mop-top viru bramboru (PMTV) ( prvního proteinu z ?triple gene block? (TGBp1)).Klon, který vykazoval nejsilnější expresi byl zkrácen o 44 aminokyselin z celkových 463.
Classification
Type
J<sub>x</sub> - Unclassified - Peer-reviewed scientific article (Jimp, Jsc and Jost)
CEP classification
EB - Genetics and molecular biology
OECD FORD branch
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Result continuities
Project
<a href="/en/project/GA522%2F04%2F1329" target="_blank" >GA522/04/1329: Molecular biological characterisation, detection and functional analysis of potato mop-top virus (PMTV)</a><br>
Continuities
Z - Vyzkumny zamer (s odkazem do CEZ)
Others
Publication year
2005
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
Protein Expression and Purification
ISSN
1046-5928
e-ISSN
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Volume of the periodical
41
Issue of the periodical within the volume
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Country of publishing house
US - UNITED STATES
Number of pages
8
Pages from-to
128-135
UT code for WoS article
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EID of the result in the Scopus database
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