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קהילה:
אסיף מאגר המחקר החקלאי
פותח על ידי קלירמאש פתרונות בע"מ -
Affinity purification of HC-Pro of potyviruses with Ni2+-NTA resin
Year:
1998
Source of publication :
Journal of Virological Methods
Authors :
גל-און, עמית
;
.
הואה, ארווה
;
.
זינגר, סימה
;
.
כדורי, דניאל
;
.
פנג, יאן הואה
;
.
רקח, בנימין
;
.
Volume :
76
Co-Authors:
Kadouri, D., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Peng, Y.-H., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Wang, Y., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Singer, S., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Huet, H., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Raccah, B., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Gal-On, A., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Facilitators :
From page:
19
To page:
29
(
Total pages:
11
)
Abstract:
The HC-Pro of zucchini yellow mosiac virus (ZYMV) was found to bind to Ni2+-NTA resin with or without His-tagging. The binding stringency was similar to that observed in proteins with a zinc finger motif like the HC-Pro. Using this characteristic we developed an efficient and rapid method (2-3 h) for purification of the HC-Pro of several potyviruses. A dominant protein of about 150 kDa was extracted and identified as the HC-Pro of ZYMV by means of immunoblotting. About 150 μg of HC-Pro were partially purified from the soluble fraction of 1 g of leaves. High titers of HC-Pro protein were obtained from plants infected with four potyviruses [ZYMV, watermelon mosaic virus II (WMVII), papaya ringspot virus (PRSV) and turnip mosaic virus (TuMV)]. The HC-Pros of potato virus Y (PVY) and tobacco vein mottling virus (TVMV) did not bind to the Ni2+-NTA resin. The ZYMV-HC-Pro purified by the Ni2+-NTA resin could bind in vitro to ZYMV virions blotted onto a membrane. All the HC-Pros which had been successfully purified by the Ni2+-NTA resin were bound in vitro to membrane-blotted ZYMV coat protein. However, only the HC-Pros of ZYMV and WMVII were able to mediate aphid transmission of purified ZYMV virions. The purification procedure described herein is efficient and convenient, and enables HC-Pro for a number of potyviruses to be obtained in larger amounts and at higher purity than possible by means of most existing methods, based on ultracentrifugation. Copyright (C) 1998 Elsevier Science B.V.
Note:
Related Files :
Animals
Brassica rapa subsp. rapa
Carica papaya
Cucurbitaceae
Cysteine Endopeptidases
Solanum tuberosum
Zucchini yellow mosaic virus
עוד תגיות
תוכן קשור
More details
DOI :
10.1016/S0166-0934(98)00119-0
Article number:
Affiliations:
Database:
סקופוס
Publication Type:
מאמר
;
.
Language:
אנגלית
Editors' remarks:
ID:
28705
Last updated date:
02/03/2022 17:27
Creation date:
17/04/2018 00:41
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Scientific Publication
Affinity purification of HC-Pro of potyviruses with Ni2+-NTA resin
76
Kadouri, D., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Peng, Y.-H., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Wang, Y., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Singer, S., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Huet, H., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Raccah, B., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Gal-On, A., Department of Virology, Agric. Res. Organisation, the V., Bet Dagan, Israel
Affinity purification of HC-Pro of potyviruses with Ni2+-NTA resin
The HC-Pro of zucchini yellow mosiac virus (ZYMV) was found to bind to Ni2+-NTA resin with or without His-tagging. The binding stringency was similar to that observed in proteins with a zinc finger motif like the HC-Pro. Using this characteristic we developed an efficient and rapid method (2-3 h) for purification of the HC-Pro of several potyviruses. A dominant protein of about 150 kDa was extracted and identified as the HC-Pro of ZYMV by means of immunoblotting. About 150 μg of HC-Pro were partially purified from the soluble fraction of 1 g of leaves. High titers of HC-Pro protein were obtained from plants infected with four potyviruses [ZYMV, watermelon mosaic virus II (WMVII), papaya ringspot virus (PRSV) and turnip mosaic virus (TuMV)]. The HC-Pros of potato virus Y (PVY) and tobacco vein mottling virus (TVMV) did not bind to the Ni2+-NTA resin. The ZYMV-HC-Pro purified by the Ni2+-NTA resin could bind in vitro to ZYMV virions blotted onto a membrane. All the HC-Pros which had been successfully purified by the Ni2+-NTA resin were bound in vitro to membrane-blotted ZYMV coat protein. However, only the HC-Pros of ZYMV and WMVII were able to mediate aphid transmission of purified ZYMV virions. The purification procedure described herein is efficient and convenient, and enables HC-Pro for a number of potyviruses to be obtained in larger amounts and at higher purity than possible by means of most existing methods, based on ultracentrifugation. Copyright (C) 1998 Elsevier Science B.V.
Scientific Publication
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