LOCUS       sequence-62727-e         863 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  natural linear DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 863)
  AUTHORS   Scholz J, Besir H, Strasser C, Suppmann S
  TITLE     A new method to customize protein expression vectors for fast, 
            efficient and background free parallel cloning.
  JOURNAL   BMC Biotechnol. 2013 Feb 14;13(1):12.
  PUBMED    23410102
REFERENCE   2  (bases 1 to 863)
  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: "BMC 
            Biotechnol."; date: "2013-02-14"; volume: "13(1)"; pages: "12"
COMMENT     Sequence Label: pCoofy1
FEATURES             Location/Qualifiers
     source          1..863
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     53..72
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        53..71
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    72..96
                     /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)."
     RBS             111..133
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             146..163
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             173..196
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     RBS             312..320
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             322..630
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MIQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSD
                     KVSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     509..528
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             650..667
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(720..738)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      734..781
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 gccggccacg atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga
       61 ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac
      121 tttaagaagg agatatacca tgaaacatca ccatcaccat cactccgcgg gtctggaagt
      181 tctgttccag gggcccatgg actaatgagg taccggatcc gaattcgagc tccgtcgaca
      241 agcttgatat cgaattcctg cagcccaaac tgcagttgac aacataaaaa ctttgtgtta
      301 tacttgtaac gtaaggaggt aatgattcag tttaaggttt acacctataa aagagagagc
      361 cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg gcgacggatg
      421 gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg
      481 gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg
      541 gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac
      601 gccattaacc tgatgttctg gggaatataa gcttgcggcc gcactcgagc accaccacca
      661 ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
      721 cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt
      781 gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta
      841 agcgcggcgg gtgtggtggt tac
//
