pLH7 [LEU2 --> HIS3 converter] (ATCC® 87545)

Applications: YI-type (integrating) shuttle vectorhost modificationmarker swap vector LEU2 --> HIS3  /  Depositors: FR Cross

Permits and Restrictions

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Designations pLH7 [LEU2 --> HIS3 converter]
Depositors FR Cross
Biosafety Level 1
Vector Information
Size (kb): 8.6999998092651370
Vector: pLH7 (phagemid)
Construction: pBC KS+
Marker(s):cmlR,kanR,HIS3
Construct size (kb): 8.699999809265137
Features: gene disruption cassette: leu2::HIS3/kanR
marker(s): cmlR
replicon: f1
replicon: pMB1
restriction site: BamHI, HpaI
restriction site: SalI, XhoI
Applications
YI-type (integrating) shuttle vector
host modification
marker swap vector LEU2 --> HIS3
Comments
Restriction digests of the clone give the following sizes (kb): BamHI/EcoRI--6.6, 2.0; XhoI--8.7.
To convert the host phenotype from LEU2 to HIS3, transform with the XhoI+HpaI digested vector and select for His+ transformants.
Some combinations of marker swap plasmids and target locus may result in relatively high reversion rates. In most but not all cases the frequencies of successful convertants are greater than 30%.
A marker swap vector designed to change the S. cerevisiae host phenotype by one-step gene disruption of the LEU2 gene with the HIS3 and kanR markers.
When swapping markers on an episomal plasmid, appropriate phenotype may result from loss of the plasmid unless a second selectable or scorable marker is used to ensure plasmid maintenance.
Vector was constructed by replacing an internal EcoRV fragment of LEU2 with a SmaI fragment containing the HIS3 and kanR coding sequences. HIS3 and LEU2 are in the same orientation.
Media ATCC Medium 1065 (see below) plus kanamycin (50.0 mcg/ml) plus chloramphenicol (25.0 mcg/ml) ATCC Medium 1065: Tryptone (Difco 0123), 10.0 g Yeast Extract (Difco 0127), 5.0 g NaCl, 10.0 g Distilled water, 1.0 L
Growth Conditions
Temperature: 37.0°C
References

Cross FR. 'Marker swap' plasmids: convenient tools for budding yeast molecular genetics. Yeast 13: 647-653, 1997. PubMed: 9200814

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