LOCUS       sequence-308949-        4504 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for the aTc-inducible expression of the Klentaq.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4504)
  AUTHORS   Milligan JN, Shroff R, Garry DJ, Ellington AD
  TITLE     Evolution of a Thermophilic Strand-Displacing Polymerase Using 
            High-Temperature Isothermal Compartmentalized Self-Replication.
  JOURNAL   Biochemistry. 2018 Aug 7;57(31):4607-4619. doi: 
            10.1021/acs.biochem.8b00200. Epub 2018 Apr 18.
  PUBMED    29629759
REFERENCE   2  (bases 1 to 4504)
  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: 
            "10.1021/acs.biochem.8b00200"; journalName: "Biochemistry"; date: 
            "2018-08-7- 7"; volume: "57"; issue: "31"; pages: "4607-4619"
COMMENT     Sequence Label: pAtetO_6xHis-Klentaq
FEATURES             Location/Qualifiers
     source          1..4504
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             14..36
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             62..79
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             80..91
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     rep_origin      complement(1957..2545)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2037..2056)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2698..3324)
                     /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
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRDNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     3229..3248
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             complement(join(3331..4122,4123..4191))
                     /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     3954..3973
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4192..4296)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4449..4504
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    4455..4473
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    4485..4503
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 aactagaaat aattttgttt aactttaaga aggagatata catatggcta gcagaggatc
       61 gcatcaccat caccatcaca tcgaagggcg ccttcttcac gagttcggac ttctcgagtc
      121 gcccaaggcc ctggaagagg cgccgtggcc tcctccagag ggggctttcg tgggatttgt
      181 gctgagtcga aaagaaccca tgtgggctga tttactcgcg ctggctgcgg caagaggggg
      241 acgtgtccat cgtgctcctg agccctataa agcactgcgc gatctcaagg aggctcgtgg
      301 cctcctcgcc aaggatcttt ctgttttggc cctgcgggag ggattggggc ttcctccggg
      361 cgacgatccc atgttacttg cgtatttatt ggaccccagt aacacgacac ctgaaggagt
      421 agcccgtcgc tatggagggg aatggaccga agaagcggga gaacgtgcgg cactgtcgga
      481 acgcctcttt gcgaacttat ggggccgtct tgaaggtgaa gagcgtcttc tgtggcttta
      541 ccgcgaagtg gagcgtccgc tgtcggcagt tctcgctcat atggaagcta cgggtgttag
      601 acttgacgtg gcatacctgc gtgcgttgtc gttagaagtg gccgaagaga ttgctagact
      661 cgaagcagaa gtttttcggt tagcgggaca tccatttaat ctcaattccc gtgaccagct
      721 ggagcgcgtt ctctttgatg aattggggct ccctgccatt gggaaaaccg aaaaaaccgg
      781 taaacgctca accagtgctg ctgtgctgga ggcactgcgt gaggcccatc cgatcgtgga
      841 gaaaattctg cagtaccggg agctcactaa gttgaaaagc acatatatcg accccctgcc
      901 ggatttaatt cacccgcgca ccggaagact ccacacaaga tttaaccaga cggcgaccgc
      961 aaccggacgc ctgtcctcat ccgatccgaa cttgcagaac atcccggtcc gtactccgtt
     1021 gggtcagcgg atccggcggg catttatcgc cgaggaaggt tggcttctgg tcgcgctgga
     1081 ctacagccaa atcgaattgc gtgtgttagc acacctttcg ggagacgaaa atttgattcg
     1141 ggtgtttcaa gagggtcgtg atattcacac ggaaaccgcg agttggatgt ttggggtgcc
     1201 tcgagaggca gtggatccac ttatgcgtcg tgcagcaaaa acaatcaatt tcggcgtcct
     1261 ctatggtatg agcgctcacc gtctttctca ggaacttgcc atcccctacg aagaggccca
     1321 ggcatttatt gagcgttatt tccagagctt tccgaaagtt agagcctgga ttgagaaaac
     1381 cttggaagaa ggtcggcgtc gggggtatgt ggagactctt tttggtcgtc ggcgctacgt
     1441 accggatctg gaagctcgtg tcaagtctgt gcgcgaagct gccgaacgta tggcatttaa
     1501 tatgccggtc caggggaccg cagcagatct tatgaagtta gcaatggtca aattgttccc
     1561 ccggcttgaa gagatggggg cgcgtatgct gcttcaggta catgatgagc ttgtcctgga
     1621 agccccgaag gaacgagcgg aagctgtggc ccgtcttgct aaagaggtca tggaaggagt
     1681 ctacccactg gccgtgccgt tggaagtgga ggtcggcatt ggagaagatt ggttatctgc
     1741 gaaagaatga taattgcctc tgccaaaatt ctgtcctcaa gcgttttagt tcgtcggtca
     1801 gtttcacctg atttacgtaa aaacccgctt cggcgggttt ttgcttttgg aggggcagaa
     1861 agatgaatga ctgtcggcca ttcgatggtg tcgggtcatg tgagcaaaag gccagcaaaa
     1921 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     1981 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2041 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2101 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2161 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2221 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2281 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2341 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2401 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     2461 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     2521 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     2581 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     2641 cacctagatc cttttaaact agtgaagtta ccatcacgga aaaaggttat gctgctttta
     2701 agacccactt tcacatttaa gttgtttttc taatccgcat atgatcaatt caaggccgaa
     2761 taagaaggct ggctctgcac cttggtgatc aaataattcg atagcttgtc gtaataatgg
     2821 cggcatacta tcagtagtag gtgtttccct ttcttcttta gcgacttgat gctcttgatc
     2881 ttccaatacg caacctaaag taaaatgccc cactgcgctg agtgcatata atgcattctc
     2941 tagtgaaaaa ccttgttggc ataaaaaggc taattgattt tcgagagttt catactgttt
     3001 ttctgtaggc cgtgtaccta aatgtacttt tgctccatcg cgatgactta gtaaagcaca
     3061 tctaaaactt ttagcgttat cacgtaaaaa atcttgccag ctttcccctt ctaaagggca
     3121 aaagtgagta tggtgcctat ctaacatctc aatggctaag gcgtcgagca aagcccgctt
     3181 attttttaca tgccaataca atgtaggctg ctctacacct agcttctggg cgagtttacg
     3241 ggttgttaaa ccttcgattc cgacctcatt aagcagctct aatgcgctgt taatcacttt
     3301 acttttatct aaacgagaca tcattaattc ctaccaatgc ttaatcagtg aggcacctat
     3361 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     3421 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagagccacg
     3481 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     3541 tggtcctgca actttatccg cctccatcca gtctatcaat tgttgccggg aagctagagt
     3601 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     3661 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     3721 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3781 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3841 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     3901 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     3961 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4021 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4081 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4141 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4201 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4261 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttcccggga actgccagac
     4321 atcaaataaa acaaaaggct cagtcggaag actgggcctt ttgttttatc tgttgtttgt
     4381 cggtgaacac tctcccgatt gcgatacaga ccctaatttc acatcatatg acactaatcc
     4441 taatttttgt tgacactcta tcattgatag agttatttta cctctcccta tcagtgatag
     4501 agaa
//
