CAT constructions with multiple unique restriction sites for the functional analysis of eukaryotic promoters and regulatory elements
Clicks: 2
ID: 291364
1987
Article Quality & Performance Metrics
Overall Quality
Not rated
Combines reader engagement with the AI quality analysis. This
article has not been analysed, so there is no overall score —
reader engagement is measured and shown alongside.
Reader Engagement
Star Article
0.3
/100
2 views
0 readers
AI Quality Assessment
Not analyzed
Readership in this journal
StarRanked #534 of 1,214 articles by views in Nucleic Acids Research
Most read
Least read
Bar heights use a square-root scale. Only the 120 most-read articles are drawn; the journal has 1,214 in total.
Mint this article as an NFT
Not yet mintedCreate a permanent, verifiable on-chain record of this article on the Scimatic Network. The NFT is held in your Journament account, and you can withdraw it to your own wallet at any time.
5
SUSD
one-off · no wallet required
Abstract
The coding region of the bacterial chloramphenicol acetyltransferase (CAT) gene is widely used as an indicator gene in gene transfer experiments dealing with regulation of transcription in eukaryotes.Chimaeric CAT fusion genes are especially useful because no endogenous CAT activity is present in eukaryotic cells and because CAT enzyme activity can be monitored by a rapid and sensitive assay (1).In order to simplify the construction of hybrid CAT genes, we have constructed the plasmids pBLCAT2 and pBLCAT3.The coding region of the CAT gene as well as the small t intron and polyadenylation signals from SV40 were inserted into the polylinker region of the high copy number plasmid pUC18 (2).Unique BgHI and Xhol restriction sites were introduced upstream of the CAT coding region by insertion of synthetic linkers.A BamHI site at the 3' end of the transcription unit was converted into a dam methylation sensitive Clal site by partial digestion with BamHI, filling -in and re -ligation.In the promoterless construction pBLCAT3 eight unique restriction sites are suitable for insertion of different eukaryotic promoters at the 5' end of the CAT gene.Four additional unique restriction sites make the insertion of regulatory signals 3' of the CAT gene possible and enable the excision of the intact fusion gene from the prokaryotic vector.The presence of die Herpes simplex virus tk promoter in pBLCAT2 permits die analysis of the effects of putative regulatory elements on a heterologous eukaryotic promoter.A BamHI/Bglll fragment from the HSV tk linker scanning mutant LS -115/ -105 (3) spanning the promoter from -105 to +51 was inserted into the corresponding restriction sites of pBLCAT3 thereby generating pBLCAT2.The modified polylinker regions at the 5' and the 3' ends have been sequenced and compiled sequences for both plasmids are available on request.
| Reference Key |
openalex_W1982598173
Use this key to autocite in the manuscript while using
SciMatic Manuscript Manager or Thesis Manager
|
|---|---|
| Authors | Bruno Luckow, Günther Schütz |
| Journal | Nucleic Acids Research |
| Year | 1987 |
| DOI |
10.1093/nar/15.13.5490
|
| URL | |
| Keywords | Keywords not found |
Citations
No citations found. To add a citation, contact the admin at info@scimatic.org
Comments
No comments yet. Be the first to comment on this article.