Improved cat gene cassette for promoter analysis and genetic constructions

Shiau, A; Smith, John L.

doi:10.1016/0378-1119(88)90406-4

Cited by 13 publications

(5 citation statements)

References 14 publications

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“…Chloramphenicol [Cm]‐resistant mutants were generated by random cartridge mutagenesis ( Labarre, Chauvat & Thuriaux 1989). A 0·8‐kb Bam HI fragment from plasmid pJS133 ( Shiau & Smith 1988) carrying the promoterless cat gene (* cat ) was ligated with fragments obtained by Sau 3A partial digestion of Synechocystis total DNA. After transformation of Synechocystis with the ligation mixture, Cm‐resistant clones were initially selected on medium C ( Kratz & Myers 1955) supplemented with 171 m M NaCl and 5 μg ml −1 Cm.…”

Section: Methodsmentioning

confidence: 99%

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios

Mikkat

Milkowski

Hagemann

2000

Plant Cell & Environment

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Section: Methodsmentioning

confidence: 99%

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios

Mikkat

Milkowski

Hagemann

2000

Plant Cell & Environment

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“…The nucleotide numbering system used is that of the parent sequence HIVNL43 (34). The CAT gene used for construction of the parent vector LTR1.4tatCAT was derived from pJS133 (45) as an EcoRI-XbaI fragment containing the CAT gene plus flanking regions. The BspEI-MscI fragment within the CAT gene was substituted with the corresponding fragment of pDS56/RBSII 6xHis (kindly provided by D. Stüber, Hoffmann-La Roche, Basel, Switzerland), a CAT expression vector carrying a silent mutation in the internal EcoRI site (GAATTC to GAATTT).…”

Section: Methodsmentioning

confidence: 99%

Exon 1 leader sequences downstream of U5 are important for efficient human immunodeficiency virus type 1 gene expression

Lenz

Scheid

Schaal

1997

J Virol

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In previous studies, little attention has been paid to maintaining the native HIV-1 leader sequence in reporter constructs analyzing the human immunodeficiency virus type 1 (HIV-1) promoter activity. To investigate a possible influence of the leader sequence on HIV-1-driven gene expression in the presence as well as in the absence of Tat, an expression vector was designed for transcripts consisting of the native HIV-1 tat 1.4 mRNA leader followed by the open reading frame for the bacterial chloramphenicol acetyltransferase (CAT). Deletion mutants with mutations within the leader sequence downstream of U5 (lsdU5) were constructed, as well as a mutant containing a mutation with a reverse orientation of this region. Quantification of CAT protein in HeLa-T4+ cells transiently transfected with wild-type and mutant leader constructs showed that the exon 1-derived lsdU5 region has an influence on basal as well as Tat-induced protein expression. The dramatic decrease in the level of CAT protein upon deletion of lsdU5 was paralleled by a drop in the steady-state level of CAT mRNA. Deletion of the exon 1-derived lsdU5 region also decreased the expression of mRNAs containing authentic HIV-1 sequences instead of CAT. The effect observed with the reporter constructs was not due to the loss of binding sites for nuclear factors, as could be shown with DBF1 and Sp1 mutant constructs. Nuclear run-on transcription assays showed that the presence or absence of lsdU5 did not influence the rate of transcription. This indicates that the exon 1 lsdU5 element functions at the posttranscriptional level in the processing, nucleocytoplasmic export, or stabilization of HIV-1 transcripts.

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“…TX302 TX337 TX529 TX530 TX701 TX705 TX709 TX717 TX725 TX726 TX729 TX764 TX768 TX769 TX771 TX773 TX778 TX779 TX780 supE44 gaIK2 rpsL31 kdgKSJ xyl-5 mtl-l rfbDI MC4100 (XpurF-lacZ)recA (Mu1+) (purR+ Kmr) MC4100 purR300 Reference 39 This study 27 Preparation of extract containing purine repressor. A 20-ml culture of strain R303 (pRRM127) was grown to late log phase in minimal medium supplemented with adenine (100 jig/ml) and kanamycin (50 ,ug/ml).…”

Section: Methodsmentioning

confidence: 99%

Genes of the Escherichia coli pur regulon are negatively controlled by a repressor-operator interaction

Shiau

Choi

et al. 1990

J Bacteriol

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Fusions of lacZ were constructed to genes in each of the loci involved in de novo synthesis of IMP. The expression of each pur-lacZ fusion was determined in isogenic purR and purR+ strains. These measurements indicated 5-to 17-fold coregulation of genes purF, purHD, purC, purMN, purL, and purEK and thus confirm the existence of a pur regulon. Gene purB, which encodes an enzyme involved in synthesis of IMP and in the AMP branch of the pathway, was not regulated by purR. Each locus of the pur regulon contains a 16-base-pair conserved operator sequence that overlaps with the promoter. The purR product, purine repressor, was shown to bind specifically to each operator. Thus, binding of repressor to each operator of pur regulon genes negatively coregulates expression.

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Improved cat gene cassette for promoter analysis and genetic constructions

Cited by 13 publications

References 14 publications

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios

Exon 1 leader sequences downstream of U5 are important for efficient human immunodeficiency virus type 1 gene expression

Genes of the Escherichia coli pur regulon are negatively controlled by a repressor-operator interaction

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Improved cat gene cassette for promoter analysis and genetic constructions

Cited by 13 publications

References 14 publications

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na+/K+ ratios

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na+/K+ ratios

Exon 1 leader sequences downstream of U5 are important for efficient human immunodeficiency virus type 1 gene expression

Genes of the Escherichia coli pur regulon are negatively controlled by a repressor-operator interaction

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The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios

The gene sll0273 of the cyanobacterium Synechocystis sp. strain PCC6803 encodes a protein essential for growth at low Na⁺/K⁺ ratios