LOCUS       sequence-165289-         878 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  E. coli expression vector derived from pET32 with Gateway cassette..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 878)
  TITLE     Olszewski lab plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 878)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET32a-DEST
FEATURES             Location/Qualifiers
     source          1..878
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        complement(join(103..109,110..127,128..133))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(158..282)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(298..312)
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     CDS             complement(328..372)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(379..396)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(406..423)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(445..771)
                     /codon_start=1
                     /gene="trxA"
                     /product="E. coli thioredoxin"
                     /label=TrxA
                     /translation="MSDKIIHLTDDSFDTDVLKADGAILVDFWAEWCGPCKMIAPILDE
                     IADEYQGKLTVAKLNIDQNPGTAPKYGIRGIPTLLLFKNGEVAATKVGALSKGQLKEFL
                     DANLA"
     RBS             779..801
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    816..840
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     complement(840..859)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(841..859)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gccattggga tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct
       61 tagctcctga aaatctcgac ggatcctaac tcaaaatcca cacattatac gagccggaag
      121 cataaagtgt aaagcctggg gtgcctaatg cggccgccat agtgactgga tatgttgtgt
      181 tttacagtat tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat
      241 atcattttac gtttctcgtt cagctttttt gtacaaactt gtgatatcag ccatggcctt
      301 gtcgtcgtcg tcggtaccca gatctgggct gtccatgtgc tggcgttcga atttagcagc
      361 agcggtttct ttcataccag aaccgcgtgg caccagacca gaagaatgat gatgatgatg
      421 gtgcatatgg ccagaaccag aaccggccag gttagcgtcg aggaactctt tcaactgacc
      481 tttagacagt gcacccactt tggttgccgc cacttcaccg tttttgaaca gcagcagagt
      541 cgggatacca cggatgccat atttcggcgc agtgccaggg ttttgatcga tgttcagttt
      601 tgcaacggtc agtttgccct gatattcgtc agcgatttca tccagaatcg gggcgatcat
      661 tttgcacgga ccgcaccact ctgcccagaa atcgacgagg atcgccccgt ccgctttgag
      721 tacatccgtg tcaaaactgt cgtcagtcag gtgaataatt ttatcgctca tatgtatatc
      781 tccttcttaa agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc
      841 cctatagtga gtcgtattaa tttcgcggga tcgagatc
//
