pMSEx-3


Vector IG Sequence Link :
General : plasmid ds-DNA 5600 BP
Functions : (cloning)
Selection : ()
Copy Number :
Hosts : (E.coli HB101)(vertebrate cells)(E.coli)
Suppliers : (ATCC)
Misc.Comments : Kanamycin resistance should be checked at regular intervals. This is best done during amplification steps using LB medium supplemented with 50 ug/ml ampicillin and 12.5 ug/ml kanamycin. (personal communication) One of 3 shuttle expression vectors (pMSEx-1-3, ATCC 37835-37837) containing a synthetic oligonucleotide providing a translation initiation signal, followed by a StuI site in 3 different positions, followed by a stop signal in all reading frames. The sequence of the oligonucleotide is: AGCTTGGTAC CATGGACGCT CGAGGCCTAG CTAGCTAGGAGCT. A EcoRI/HindIII digest gives fragments of 2.63, 1.68, 0.97 and 0.32 kb. (personal communication) The SV40 promoter has a 3 bp deletion at the BglI site within the origin of replication to prevent autonomous replication in the presence of SV40 large T-antigen. The MSV LTR provides a polyadenylation signal as well as an enhancer for the P1/thymidine kinase promoter regulating the Tn5 kanR/G418R transcription unit. Restriction digests of the clone give the following sizes (kb): HindIII--3.2, 2.9; BglII--6.1; EcoRI/HindIII--2.8, 1.8, 1.0, 0.4; EcoRI--4.7, 1.4; consistent with deposited DNA. (ATCC staff) The order of the major features in this plasmid is:SV40 promoter - oligonucleotide/StuI/oligonucleotide - MLV-LTR - P1 promoter - TK promoter - kanR/G418R - TK polyadenylation signal - ampR. Medium is 1227 LB plus ampicillin.
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