LOCUS       sequence-437767-        4955 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Testing the activity of LguI-associated switch methylases.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4955)
  AUTHORS   Flores-Fernandez CN, Lin D, Robins K, O'Callaghan CA
  TITLE     DNA methylases for site-selective inhibition of type IIS restriction
            enzyme activity.
  JOURNAL   Appl Microbiol Biotechnol. 2024 Jan 25;108(1):174. doi: 
            10.1007/s00253-024-13015-7.
  PUBMED    38270650
REFERENCE   2  (bases 1 to 4955)
  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: "Appl 
            Microbiol Biotechnol."; date: "2024-01-25"; volume: "108(1):174. 
            doi"; pages: " 10.1007/s00253-024-13015-7"
COMMENT     Sequence Label: POC1401
FEATURES             Location/Qualifiers
     source          1..4955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    89..110
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(125..130,131..148,149..155)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    163..179
                     /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     168..190
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     187..203
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     187..203
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             199..522
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     misc_feature    216..272
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(273..290)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(273..289)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(282..304)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(498..517)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     misc_feature    661..685
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(841..1989)
                     /codon_start=1
                     /product="trans-acting replication protein that binds to 
                     and activates oriV"
                     /label=trfA
                     /translation="MNRTFDRKAYRQELIDAGFSAEDAETIASRTVMRAPRETFQSVGS
                     MVQQATAKIERDSVQLAPPALPAPSAAVERSRRLEQEAAGLAKSMTIDTRGTMTTKKRK
                     TAGEDLAKQVSEAKQAALLKHTKQQIKEMQLSLFDIAPWPDTMRAMPNDTARSALFTTR
                     NKKIPREALQNKVIFHVNKDVKITYTGVELRADDDELVWQQVLEYAKRTPIGEPITFTF
                     YELCQDLGWSINGRYYTKAEECLSRLQATAMGFTSDRVGHLESVSLLHRFRVLDRGKKT
                     SRCQVLIDEEIVVLFAGDHYTKFIWEKYRKLSPTARRMFDYFSSHREPYPLKLETFRLM
                     CGSDSTRVKKWREQVGEACEELRGSGLVEHAWVNDDLVHCKR"
     rep_origin      complement(2325..2913)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2405..2424)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3084..3875,3876..3944))
                     /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     3707..3726
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3945..4049)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4136..4767
                     /label=oriV
                     /note="incP origin of replication"
     misc_feature    4859..4883
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 cactctgtga agctcttcag cctgaagagc ttcgcagctg gcacgacagg tttcccgact
       61 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat taggcacccc
      121 aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc ggataacaat
      181 ttcacacagg aaacagctat gaccatgatt acgccaagct tgcatgcctg caggtcgact
      241 ctagaggatc cccgggtacc gagctcgaat tcactggccg tcgttttaca acgtcgtgac
      301 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
      361 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
      421 ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
      481 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
      541 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
      601 caagctgtga cttgtcttct gagacctgaa gagcacgaag tggtttaaac tatcagtgtt
      661 tgacaggata tattggcggg taaacctaag agaaaagagc gtttattaga ataatcggat
      721 atttaaaagg gcgtgaaaag gtttatccgt tcgtccattt gtatgtgcat gccaaccaca
      781 gggttcccca gatcaggcgc tggctgctga acccccagcc ggaactgacc ccacaaggcc
      841 ctagcgtttg caatgcacca ggtcatcatt gacccaggcg tgttccacca ggccgctgcc
      901 tcgcaactct tcgcaggctt cgccgacctg ctcgcgccac ttcttcacgc gggtggaatc
      961 cgatccgcac atgaggcgga aggtttccag cttgagcggg tacggctccc ggtgcgagct
     1021 gaaatagtcg aacatccgtc gggccgtcgg cgacagcttg cggtacttct cccatatgaa
     1081 tttcgtgtag tggtcgccag caaacagcac gacgatttcc tcgtcgatca ggacctggca
     1141 acgggacgtt ttcttgccac ggtccaggac gcggaagcgg tgcagcagcg acaccgattc
     1201 caggtgccca acgcggtcgg acgtgaagcc catcgccgtc gcctgtaggc gcgacaggca
     1261 ttcctcggcc ttcgtgtaat accggccatt gatcgaccag cccaggtcct ggcaaagctc
     1321 gtagaacgtg aaggtgatcg gctcgccgat aggggtgcgc ttcgcgtact ccaacacctg
     1381 ctgccacacc agttcgtcat cgtcggcccg cagctcgacg ccggtgtagg tgatcttcac
     1441 gtccttgttg acgtggaaaa tgaccttgtt ttgcagcgcc tcgcgcggga ttttcttgtt
     1501 gcgcgtggtg aacagggcag agcgggccgt gtcgtttggc atcgctcgca tcgtgtccgg
     1561 ccacggcgca atatcgaaca aggaaagctg catttccttg atctgctgct tcgtgtgttt
     1621 cagcaacgcg gcctgcttgg cctcgctgac ctgttttgcc aggtcctcgc cggcggtttt
     1681 tcgcttcttg gtcgtcatag ttcctcgcgt gtcgatggtc atcgacttcg ccaaacctgc
     1741 cgcctcctgt tcaagacgac gcgaacgctc cacggcggcc gatggcgcgg gcagggcagg
     1801 gggagccagt tgcacgctgt cgcgctcgat cttggccgta gcttgctgga ccatcgagcc
     1861 gacggactgg aaggtttcgc ggggcgcacg catgacggtg cggcttgcga tggtttcggc
     1921 atcctcggcg gaaaaccccg cgtcgatcag ttcttgcctg tatgccttcc ggtcaaacgt
     1981 ccgattcatt caccctcctt gcgggattgc cccgactcac gccggggcaa tgtgccctta
     2041 ttcctgattt gacccgcctg gtgccttggt gtccagataa tccaccttat cggcaatgaa
     2101 gtcggtcccg tagaccgtct ggccgtcctt ctcgtacttg gtattccgaa tcttgccctg
     2161 cacgaatacc agcgacccct tgcccaaata cttgccgtgg gcctcggcct gagagccaaa
     2221 acacttgatg cggaagaagt cggtgcgctc ctgcttgtcg ccggcatcgt tgcgccacat
     2281 ctaggatctg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     2341 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     2401 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     2461 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     2521 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     2581 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     2641 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     2701 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     2761 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     2821 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     2881 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     2941 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     3001 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     3061 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     3121 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     3181 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaacc acgctcaccg
     3241 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     3301 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     3361 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     3421 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     3481 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     3541 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     3601 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     3661 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     3721 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     3781 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     3841 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     3901 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     3961 tattattgaa gcatttatca aggttattgt ctcatgagcg gatacatatt tgaatgtatt
     4021 tagaaaaata aacaaatagg ggttccgcgc acgaattggc cagcgctgcc atttttgggg
     4081 tgaggccgtt cgcggccgag gggcgcagcc cctgggggga tgggaggccc gcgttagcgg
     4141 gccgggaggg ttcgagaagg gggggcaccc cccttcggcg tgcgcggtca cgcgcacagg
     4201 gcgcagccct ggttaaaaac aaggtttata aatattggtt taaaagcagg ttaaaagaca
     4261 ggttagcggt ggccgaaaaa cgggcggaaa cccttgcaaa tgctggattt tctgcctgtg
     4321 gacagcccct caaatgtcaa taggtgcgcc cctcatctgt cagcactctg cccctcaagt
     4381 gtcaaggatc gcgcccctca tctgtcagta gtcgcgcccc tcaagtgtca ataccgcagg
     4441 gcacttatcc ccaggcttgt ccacatcatc tgtgggaaac tcgcgtaaaa tcaggcgttt
     4501 tcgccgattt gcgaggctgg ccagctccac gtcgccggcc gaaatcgagc ctgcccctca
     4561 tctgtcaacg ccgcgccggg tgagtcggcc cctcaagtgt caacgtccgc ccctcatctg
     4621 tcagtgaggg ccaagttttc cgcgaggtat ccacaacgcc ggcggccgcg gtgtctcgca
     4681 cacggcttcg acggcgtttc tggcgcgttt gcagggccat agacggccgc cagcccagcg
     4741 gcgagggcaa ccagcccggt gagcgtcgca aaggagatcc tgatctgact gatgggctgc
     4801 ctgtatcgag tggtgatttt gtgccgagct gccggtcggg gagctgttgg ctggctggtg
     4861 gcaggatata ttgtggtgta aacaaattga cgcttagaca acttaataac acattgcgga
     4921 cgtttttaat gtactggggt ggatgcagtg ggccc
//
