pATH20 (ATCC® 37700)

Applications: expression vectorvector permitting construction of fusion proteins  /  Depositors: A Tzagoloff, TJ Koerner

Permits and Restrictions

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Designations pATH20
Depositors A Tzagoloff, TJ Koerner
Biosafety Level 1
Vector Information
Size (kb): 3.8499999046325680
Vector: pATH20 (plasmid)
Promoters: Promoter trp
Construction: pBR322, polylinker, trp promoter, trpE
Marker(s):ampR
Construct size (kb): 3.849999904632568
Features: marker(s): ampR
promoter: trp
replicon: pMB1
Applications
expression vector
vector permitting construction of fusion proteins
Comments
The nucleotides remaining in the codon after cutting the vector with the given enzyme are: BamHI-1, ClaI-?, EcoRI-1, HindIII-1, KpnI-2, PstI-2, SacI-2, SalI-1, SmaI-1, SphI-2, XbaI-1, XmaI-2.
Restriction digests of the clone give the following sizes (kb): BamHI--3.85; EcoRI--3.85; HindIII--3.85.
To prevent recircularization of the vector, the depositors recommend including 30 - 50 U of calf alkaline phosphatase in the restriction enzyme digestion used to prepare the vector.
Fusion proteins are produced in Escherichia coli usually in an insoluble form which facilitates purification. Hybrid proteins larger than 90 kDa are not expressed as efficiently as shorter ones.
Escherichia coli containing pATH vectors should be grown on medium supplemented with tryptophan to prevent expression. Storage as DNA at -20C is preferred; long term storage as cells may result in selection of non-expressing promoter mutations.
One of a series of vectors (ATCC 37695-37703) designed to facilitate expression of an open reading frame as a trpE fusion protein.
Nucleotide 1 is the first in the trp fragment, at the beginning of a PvuII site 261 nucleotides upstream of the transcription start site. The multiple cloning site follows codon 323 of trpE.
Media 1065 plus ampicillin (50 mcg/ml) and tryptophan (10 mcg/ml)
Growth Conditions
Temperature: 37.0°C
References

Koerner TJ, et al. High-expression vectors with multiple cloning sites for construction of trpE-fusion genes: pATH vectors. Methods Enzymol. 194: 477-490, 1991. PubMed: 2005804

Shipped freeze-dried
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