Plasmid 37060: pHSS6

• Gene/insert name: None
• Vector backbone: pMB8
  (Search Vector Database)
• Vector type: Cloning vector
• Backbone size (bp): 2320
• Modifications to Backbone: This Tn3-free vector was designed to carry a minimal amount of nonessential information to favor transposition into cloned DNA sequences rather than into the vector. The important part of the vector is the polylinker surrounded by restriction endonuclease Not I sites. Since Not I has an 8-bp recognition sequence (5'-GCGGCCGC) it would be predicted to cut once every 16 kilobases (kb) in a random DNA sequence. In an A-T-rich organism like yeast, the enzyme has a probability of cutting every 390 kb (assuming 60% AT). Therefore, DNA fragments that have been cloned between the Not I sites have a high probability of being excised intact by Not I. The basic vector used in the procedure is derived from an amplifiable pMB8 replicon (pFH97; Heffron et al., 1978), the kanamycin resistance determinant (Kmr) from Tn5, and a synthetic double-stranded DNA molecule containing recognition sites for the restriction enzymes Not I, BamHI, and EcoRI. The vectors also contain the polylinker from pLink322 (Maniatis et al., 1982) that contains sites for Cla I, HindIII, Xba I, Bgl II, and Pst I. The vector is 2.3 kb in size and contains less than 1000 bp of nonessential DNA sequence.
• 5' sequencing primer: NA
• 3' sequencing primer: Kan-R
• Bacterial resistance(s) Kanamycin
• Growth strain(s) DH5alpha
• Growth temperature (℃): 37
• High or low copy: Unknown
• Sequence: View sequences (3)
• Map: View map
• Principal Investigator: Fred Heffron
• Principal Investigator: Hank Seifert
• Terms and Licenses: MTA

Article: Shuttle mutagenesis: a method of transposon mutagenesis for Saccharomyces cerevisiae. Seifert et al (Proc Natl Acad Sci U S A. 1986 Feb;83(3):735-9. PubMed)