LOCUS       sequence-205218-        4709 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses WspR WT under a T7 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4709)
  AUTHORS   Wang XC, Wilson SC, Hammond MC
  TITLE     Next-generation RNA-based fluorescent biosensors enable anaerobic 
            detection of cyclic di-GMP.
  JOURNAL   Nucleic Acids Res. 2016 Jul 5. pii: gkw580.
  PUBMED    27382070
REFERENCE   2  (bases 1 to 4709)
  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: "Nucleic 
            Acids Res. 2016 Jul 5. pii: gkw580."
COMMENT     Sequence Label: pCOLA-T7-WspR
FEATURES             Location/Qualifiers
     source          1..4709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    8..32
                     /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)."
     CDS             88..105
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        219..237
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    238..262
                     /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)."
     CDS             1361..1405
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(1443..1461)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1457..1504
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(1734..2549)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1734..2549)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2457..2476
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(2550..2641)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2659..3294)
                     /direction=LEFT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication 
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     primer_bind     complement(2804..2823)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3492..4574)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4536..4555
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4575..4652)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(4698..4709,1..7)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ctatagggga attgtgagcg gataacaatt cccctgtaga aataattttg tttaacttta
       61 ataaggagat ataccatggg cagcagccat caccatcatc accacagcca ggatccgaat
      121 tcgagctcgg cgcgcctgca ggtcgacaag cttgcggccg cataatgctt aagtcgaaca
      181 gaaagtaatc gtattgtaca cggccgcata atcgaaatta atacgactca ctatagggga
      241 attgtgagcg gataacaatt ccccatctta gtatattagt taagtataag aaggagatat
      301 acatatgcac aaccctcatg agagcaagac cgacctgggc gcccctctgg acggcgcggt
      361 catggtactg cttgtcgatg atcaggccat gatcggagag gctgtgcgtc gctcactggc
      421 gagcgaggcg ggcatcgact tccatttctg ttccgatccg cagcaggcgg tggcggtggc
      481 caaccagatc aagccgacgg tgatcctcca ggacctggtg atgcccggcg tcgacggcct
      541 cacgctgctc gctgcctacc gcggcaaccc ggcgacccgc gacatcccga tcatcgtcct
      601 gtcgaccaag gaagagccga cggtgaagag cgcggcgttc gccgccggcg ccaacgacta
      661 cctggtcaag ctgccggacg ccatcgagct ggtggcgcgg atccgctacc actcgcgttc
      721 gtacatcgcc ctgcagcaac gcgacgaggc ctatcgggcg ctgcgcgaga gccagcagca
      781 gttgctggag accaacctgg tgctgcagcg gctgatgaac tccgacggcc tgaccgggct
      841 ctccaaccgt cgtcacttcg acgaatacct ggagatggag tggcgccgct cgctgcgcga
      901 gcagtcgcaa ctgtcgctgc tgatgatcga cgtggactat ttcaagagct acaacgacac
      961 cttcggccac gtcgccggcg acgaggccct gcgccaggtc gccggggcca tccgcgaggg
     1021 ctgcagtcgc tcctcggacc tggcggctcg gtacggcggc gaggagttcg ccatggtcct
     1081 cccgggcacc tcgccgggcg gcgcacggtt gctggcggag aaggtccggc gcacggtgga
     1141 gagcctgcag atcagccacg accagccacg cccgggctcg cacctgaccg tcagcatcgg
     1201 cgtcagcacc ctggtgccgg gcggcggcgg gcaaaccttc cgggttctca tcgagatggc
     1261 cgaccaggcg ctctaccagg ccaagaacaa tggccgcaac caggtgggcc tgatggaaca
     1321 gccggtgccg ccggccccgg cgggctgact cgagtctggt aaagaaaccg ctgctgcgaa
     1381 atttgaacgc cagcacatgg actcgtctac tagcgcagct taattaacct aggctgctgc
     1441 caccgctgag caataactag cataacccct tggggcctct aaacgggtct tgaggggttt
     1501 tttgctgaaa cctcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa
     1561 accggtaaac cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac
     1621 aagctgacga ccgggtctcc gcaagtggca cttttcgggg aaatgtgcgc ggaaccccta
     1681 tttgtttatt tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa
     1741 aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat
     1801 ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg
     1861 gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat
     1921 ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc
     1981 ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta
     2041 cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga
     2101 gcgagacgaa atacgcggtc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac
     2161 cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct
     2221 aatacctgga atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga
     2281 gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg
     2341 accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct
     2401 ggcgcatcgg gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg
     2461 cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag
     2521 caagacgttt cccgttgaat atggctcata ctcttccttt ttcaatatta ttgaagcatt
     2581 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     2641 ataggcatgc tagcgcagaa acgtcctaga agatgccagg aggatactta gcagagagac
     2701 aataaggccg gagcgaagcc gtttttccat aggctccgcc cccctgacga acatcacgaa
     2761 atctgacgct caaatcagtg gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2821 ccccctgatg gctccctctt gcgctctcct gttcccgtcc tgcggcgtcc gtgttgtggt
     2881 ggaggcttta cccaaatcac cacgtcccgt tccgtgtaga cagttcgctc caagctgggc
     2941 tgtgtgcaag aaccccccgt tcagcccgac tgctgcgcct tatccggtaa ctatcatctt
     3001 gagtccaacc cggaaagaca cgacaaaacg ccactggcag cagccattgg taactgagaa
     3061 ttagtggatt tagatatcga gagtcttgaa gtggtggcct aacagaggct acactgaaag
     3121 gacagtattt ggtatctgcg ctccactaaa gccagttacc aggttaagca gttccccaac
     3181 tgacttaacc ttcgatcaaa ccgcctcccc aggcggtttt ttcgtttaca gagcaggaga
     3241 ttacgacgat cgtaaaagga tctcaagaag atcctttacg gattcccgac accatcactc
     3301 tagatttcag tgcaatttat ctcttcaaat gtagcacctg aagtcagccc catacgatat
     3361 aagttgtaat tctcatgtta gtcatgcccc gcgcccaccg gaaggagctg actgggttga
     3421 aggctctcaa gggcatcggt cgagatcccg gtgcctaatg agtgagctaa cttacattaa
     3481 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     3541 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgccagggt ggtttttctt
     3601 ttcaccagtg agacgggcaa cagctgattg cccttcaccg cctggccctg agagagttgc
     3661 agcaagcggt ccacgctggt ttgccccagc aggcgaaaat cctgtttgat ggtggttaac
     3721 ggcgggatat aacatgagct gtcttcggta tcgtcgtatc ccactaccga gatgtccgca
     3781 ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg
     3841 gcaaccagca tcgcagtggg aacgatgccc tcattcagca tttgcatggt ttgttgaaaa
     3901 ccggacatgg cactccagtc gccttcccgt tccgctatcg gctgaatttg attgcgagtg
     3961 agatatttat gccagccagc cagacgcaga cgcgccgaga cagaacttaa tgggcccgct
     4021 aacagcgcga tttgctggtg acccaatgcg accagatgct ccacgcccag tcgcgtaccg
     4081 tcttcatggg agaaaataat actgttgatg ggtgtctggt cagagacatc aagaaataac
     4141 gccggaacat tagtgcaggc agcttccaca gcaatggcat cctggtcatc cagcggatag
     4201 ttaatgatca gcccactgac gcgttgcgcg agaagattgt gcaccgccgc tttacaggct
     4261 tcgacgccgc ttcgttctac catcgacacc accacgctgg cacccagttg atcggcgcga
     4321 gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg
     4381 ccaatcagca acgactgttt gcccgccagt tgttgtgcca cgcggttggg aatgtaattc
     4441 agctccgcca tcgccgcttc cactttttcc cgcgttttcg cagaaacgtg gctggcctgg
     4501 ttcaccacgc gggaaacggt ctgataagag acaccggcat actctgcgac atcgtataac
     4561 gttactggtt tcacattcac caccctgaat tgactctctt ccgggcgcta tcatgccata
     4621 ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct cgacgctctc ccttatgcga
     4681 ctcctgcatt aggaaattaa tacgactca
//
