LOCUS       sequence-417772-        3980 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses 2',3'-cyclic nucleotide phosphodiesterase catalytic 
            domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3980)
  AUTHORS   Fontaine BM, Martin KS, Garcia-Rodriguez JM, Jung C, Briggs L, 
            Southwell JE, Jia X, Weinert EE
  TITLE     RNase I regulates Escherichia coli 2',3'-cyclic nucleotide 
            monophosphate levels and biofilm formation.
  JOURNAL   Biochem J. 2018 Apr 30;475(8):1491-1506. doi: 10.1042/BCJ20170906.
  PUBMED    29555843
REFERENCE   2  (bases 1 to 3980)
  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"; doi: "10"; journalName: 
            "Biochem J"; date: "2018-04-30- 30"; volume: "475"; issue: "8"; 
            pages: "1491-1506"
COMMENT     Sequence Label: pKT-CNP
FEATURES             Location/Qualifiers
     source          1..3980
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(541..559)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(566..583)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(566..582)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(575..597)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      724..1179
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(811..830)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1021..1042
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1205..1309
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1310..1378,1379..2170)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1528..1547)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2341..2929
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2830..2849
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2991..3617)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     3522..3541
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        3633..3688
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    3639..3657
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    3669..3687
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        3712..3730
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             3762..3784
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 tttggcaaac gcccgccggg tgttctgcat tgcaccacga aattctgtga ttatggcaaa
       61 gcgaccggtg ccgaagaata cgcccagcaa gatgtggttc gtcgctctta cggcaaagca
      121 tttaaactgt ccatctcagc tctgttcgtg accccgaaaa cggcaggtgc tcaggtcgtg
      181 ctgaatgaac aggaactgca actgtggccg agcgatctgg acaaaccgag ctctagtgaa
      241 agcctgccgc cgggcagccg tgcgcatgtt accctgggtt gtgccgcgga tgttcagccg
      301 gtccaaacgg gcctggacct gctggaaatt ctgcagcaag tgaaaggcgg ttcccagggc
      361 gaagaagttg gtgaactgcc gcgtggtaaa ctgtatagcc tgggcaaagg tcgctggatg
      421 ctgtccctgg caaagaaaat ggaagtgaaa gctatcttta cgggctacta cggctgaact
      481 agtgcggccg cttgataaag cccgcctaat gagcgggctt ttttttgagc tccaattcgc
      541 cctatagtga gtcgtattac aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa
      601 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
      661 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
      721 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
      781 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      841 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      901 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      961 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     1021 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     1081 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     1141 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt ttcggggaaa
     1201 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1261 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1321 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     1381 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     1441 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     1501 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     1561 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     1621 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     1681 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     1741 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     1801 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     1861 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     1921 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     1981 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     2041 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     2101 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     2161 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     2221 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     2281 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     2341 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     2401 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     2461 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     2521 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     2581 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     2641 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     2701 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     2761 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     2821 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     2881 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     2941 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgc ttaagaccca
     3001 ctttcacatt taagttgttt ttctaatccg caaatgatca attcaaggcc gaataagaag
     3061 gctggctctg caccttggtg atcaaataat tcgatagctt gtcgtaataa tggcggcata
     3121 ctatcagtag taggtgtttc cctttcttct ttagcgactt gatgctcttg atcttccaat
     3181 acgcaaccta aagtaaaatg ccccacagcg ctgagtgcat ataatgcatt ctctagtgaa
     3241 aaaccttgtt ggcataaaaa ggctaattga ttttcgagag tttcatactg tttttctgta
     3301 ggccgtgtac ctaaatgtac ttttgctcca tcgcgatgac ttagtaaagc acatctaaaa
     3361 cttttagcgt tattacgtaa aaaatcttgc cagctttccc cttctaaagg gcaaaagtga
     3421 gtatggtgcc tatctaacat ctcaatggct aaggcgtcga gcaaagcccg cttatttttt
     3481 acatgccaat acaatgtagg ctgctctaca cctagcttct gggcgagttt acgggttgtt
     3541 aaaccttcga ttccgacctc attaagcagc tctaatgcgc tgttaatcac tttactttta
     3601 tctaatctcg acatcattaa ttcctaattt ttgttgacac tctatcattg atagagttat
     3661 tttaccactc cctatcagtg atagagaaaa gtctagccga tcccgcgaaa ttaatacgac
     3721 tcactatagg gagaccacaa cggtttccct ctagaaataa ttttgtttaa ctttaagaag
     3781 gagatataca tatgggcctg gaaaaagatt tcctgccgct gtactttggc tggttcctga
     3841 cgaaaaaatc gagcgaaacc ctgcgtaaag ctggtcaagt gttcctggaa gaactgggca
     3901 accataaagc gtttaagaaa gaactgcgtc acttcatttc tggtgatgaa ccgaaagaaa
     3961 aactggacct ggtcagctat
//
