Abstract:
:Our goal was to develop a system to study proteins that associate in vivo with the Moloney murine leukemia virus (M-MuLV) enhancer elements by the isolation of intact proviral chromatin. The M-MuLV long terminal repeats (LTRs) contain tandemly repeated transcriptional enhancer sequences consisting of smaller motifs that bind cellular DNA-binding proteins implicated in transcriptional regulation. The M-MuLV enhancers are also important for disease specificity and latency of disease induction. To enrich for proviral chromatin containing M-MuLV LTR sequences, an affinity purification scheme was employed that relies on the affinity of bacterial Lac repressor protein for Lac operator (LacO) DNA sequences. An infectious M-MuLV recombinant was constructed that contains bacterial LacO sequences inserted into a nonessential region downstream from the 5' LTR of the virus (M-MuLV-LacO). Nuclei from M-MuLV-LacO-infected cells were digested with PvuII (which will liberate an LTR fragment containing LacO sequences), and digested chromatin was leached from the nuclei in hypotonic buffer. M-MuLV-LacO chromatin was then recovered by binding to an affinity matrix consisting of a beta-galactosidase-Lac repressor fusion protein anchored to acrylamide beads by an anti-beta-galactosidase monoclonal antibody [7]. Specifically bound chromatin was eluted under physiological conditions by incubation with the galactose analog isopropyl-beta-D-thiogalactopyranoside. Southern blot analysis confirmed the specific enrichment of M-MuLV proviral chromatin by this method.
journal_name
J Biomed Scijournal_title
Journal of biomedical scienceauthors
Granger SW,Fan Hdoi
10.1007/BF02256602keywords:
subject
Has Abstractpub_date
2001-05-01 00:00:00pages
278-89issue
3eissn
1021-7770issn
1423-0127pii
54044journal_volume
8pub_type
杂志文章abstract:BACKGROUND:Breast cancer is considered as an increasing major life-threatening concern among the malignancies encountered globally in females. Traditional therapy is far from satisfactory due to drug resistance and various side effects, thus a search for complementary/alternative medicines from natural sources with les...
journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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更新日期:2013-05-31 00:00:00
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
pub_type: 杂志文章,评审
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更新日期:2019-10-20 00:00:00
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journal_title:Journal of biomedical science
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更新日期:2015-08-08 00:00:00
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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更新日期:1998-01-01 00:00:00
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journal_title:Journal of biomedical science
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doi:10.1186/1423-0127-17-63
更新日期:2010-07-28 00:00:00
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更新日期:2019-10-21 00:00:00
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journal_title:Journal of biomedical science
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更新日期:2005-12-01 00:00:00
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journal_title:Journal of biomedical science
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更新日期:2017-02-07 00:00:00
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更新日期:2006-07-01 00:00:00
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journal_title:Journal of biomedical science
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更新日期:2001-09-01 00:00:00
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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journal_title:Journal of biomedical science
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