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Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions.
Gene 1986; 45(3):299-310GENE

Abstract

We report yeast/Escherichia coli shuttle vectors suitable for fusing yeast promoter and coding sequences to the lacZ gene of E. coli. The vectors contain a region of multiple unique restriction sites including EcoRI, KpnI, SmaI, BamHI, XbaI, SalI, PstI, SphI and HindIII. The region with the unique cloning sites has been introduced in both orientations with respect to lacZ and occurs proximal to the eighth codon of the gene. All the restriction sites have been phased to three different reading frames. Two series of vectors have been constructed. The first series (YEp) has two origins of replication (ori), i.e., of the yeast 2 mu circle and of the ColE1 plasmid of E. coli, and can therefore replicate autonomously in both organisms. These shuttle vectors also have the ApR gene of E. coli and either the yeast LEU2 or URA3 genes to allow for selection of both E. coli and yeast transformants. The second series of vectors (YIp) are identical in all respects to the YEp vectors except that they lack the 2 mu ori. The YIp vectors can be used to integrate lacZ fusions into yeast chromosomal DNA. None of the vectors express beta-galactosidase (beta Gal) in yeast or E. coli in the absence of inserted yeast promoter sequences. The 5'-nontranslated sequences and parts of the coding sequences of various yeast genes have been cloned into representative lacZ fusion vectors. In-frame gene fusions can be detected by beta Gal activity when either yeast or E. coli clones are plated on media containing XGal indicator. Quantitative determinations of promoter activity were made by colorimetric assay of beta Gal activity in whole cells. Fusion of the yeast CYC1 gene to lacZ in one of the vectors allowed detection of regulated expression of this gene when cells were grown under conditions of catabolite repression or derepression.

Authors

No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

3026915

Citation

Myers, A M., et al. "Yeast Shuttle and Integrative Vectors With Multiple Cloning Sites Suitable for Construction of lacZ Fusions." Gene, vol. 45, no. 3, 1986, pp. 299-310.
Myers AM, Tzagoloff A, Kinney DM, et al. Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions. Gene. 1986;45(3):299-310.
Myers, A. M., Tzagoloff, A., Kinney, D. M., & Lusty, C. J. (1986). Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions. Gene, 45(3), pp. 299-310.
Myers AM, et al. Yeast Shuttle and Integrative Vectors With Multiple Cloning Sites Suitable for Construction of lacZ Fusions. Gene. 1986;45(3):299-310. PubMed PMID: 3026915.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Yeast shuttle and integrative vectors with multiple cloning sites suitable for construction of lacZ fusions. AU - Myers,A M, AU - Tzagoloff,A, AU - Kinney,D M, AU - Lusty,C J, PY - 1986/1/1/pubmed PY - 1986/1/1/medline PY - 1986/1/1/entrez SP - 299 EP - 310 JF - Gene JO - Gene VL - 45 IS - 3 N2 - We report yeast/Escherichia coli shuttle vectors suitable for fusing yeast promoter and coding sequences to the lacZ gene of E. coli. The vectors contain a region of multiple unique restriction sites including EcoRI, KpnI, SmaI, BamHI, XbaI, SalI, PstI, SphI and HindIII. The region with the unique cloning sites has been introduced in both orientations with respect to lacZ and occurs proximal to the eighth codon of the gene. All the restriction sites have been phased to three different reading frames. Two series of vectors have been constructed. The first series (YEp) has two origins of replication (ori), i.e., of the yeast 2 mu circle and of the ColE1 plasmid of E. coli, and can therefore replicate autonomously in both organisms. These shuttle vectors also have the ApR gene of E. coli and either the yeast LEU2 or URA3 genes to allow for selection of both E. coli and yeast transformants. The second series of vectors (YIp) are identical in all respects to the YEp vectors except that they lack the 2 mu ori. The YIp vectors can be used to integrate lacZ fusions into yeast chromosomal DNA. None of the vectors express beta-galactosidase (beta Gal) in yeast or E. coli in the absence of inserted yeast promoter sequences. The 5'-nontranslated sequences and parts of the coding sequences of various yeast genes have been cloned into representative lacZ fusion vectors. In-frame gene fusions can be detected by beta Gal activity when either yeast or E. coli clones are plated on media containing XGal indicator. Quantitative determinations of promoter activity were made by colorimetric assay of beta Gal activity in whole cells. Fusion of the yeast CYC1 gene to lacZ in one of the vectors allowed detection of regulated expression of this gene when cells were grown under conditions of catabolite repression or derepression. SN - 0378-1119 UR - https://www.unboundmedicine.com/medline/citation/3026915/Yeast_shuttle_and_integrative_vectors_with_multiple_cloning_sites_suitable_for_construction_of_lacZ_fusions_ L2 - https://linkinghub.elsevier.com/retrieve/pii/0378-1119(86)90028-4 DB - PRIME DP - Unbound Medicine ER -