LOCUS       sequence-439148-        7104 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LguI-associated switch methylase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  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 7104)
  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: POC1482
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2045..2067
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2082..2106
                     /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     complement(2106..2125)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2107..2125)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2209..2228)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2434..2511
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2512..3594
                     /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"
     CDS             4403..4594
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4588..4610)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4696..4835
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4751..4770
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4850..4867)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5021..5609)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5101..5120)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5720..5739)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5731..6546
                     /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             5731..6546
                     /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     complement(5804..5823)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(6638..7093)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6775..6796)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6987..7006
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttcc
      181 cggcgcgcaa accatcgctt aacttgttcg cgatactctg ttgtaaacgt gcgatctcgt
      241 tcaggcggtc atttaaggct acctcttcga ccagcaattc ctgcacctca cgccagtcat
      301 cggaagcatc cagcactgat gtagggacca tcagagccgc aagatcatct gcgtgtaagt
      361 gcgccgtctg accgcgcacc atttcctgga tctgagaggc cactgtcggt tggcgtaagt
      421 aggccagcag taacataggt gcaatgcgtt cttgctttgg gcgcaccaac attacttctc
      481 ccacaaatga tgcacggccg cctacccatt ctggaacttt ggtgataatg tcaactttct
      541 ttgcaatata gactggactg tgtgcgctac tagttaagac gatgtcgccg tttttgacca
      601 attgaatgcc cccaagtttg cgactagtca tcgttgacgt atcaataaag ttacggtcgc
      661 gagcaatcca gttaattcca ctcccagtca agttgcctac cttgataaga aaaagaccgc
      721 cattctcact atatgctgca cgaggtggcg tacgtccggt gcgaatctcg ctggccaaat
      781 cacctaaagg aacaccaagg ttcgagcgac gttttgacgc ggacttcgca gacagtgcct
      841 cgaaggtgcg gtcagccttc tggggagcca ggacagaaac ataatcacag ggtgaacctt
      901 ttgggtcgcg catatgcttg gctaaaccat caagttgcga cccttgacgc gggcgtcctg
      961 tgctgtcgta tccgacattc gataaggtcg cctggactac tcctactttc ttttcacggt
     1021 ccttatcaga agcgtatttc tcaatgaagg ctaccactgt gtttgtctgc gccccggtca
     1081 ccaagaacgt ttccggtggc aattgaatga cagccttcag gtagccaagt ttcgccaaca
     1141 agctacgttc gtacgctgac gatgggttcg tcagaacgct gcgaggcaac actacagcca
     1201 aagttccgcc aggcttcagc cagcgaagcg attgttcaac aaataaccat tctgagcttt
     1261 ggcgggcgcg tggctctcca tttgctgtaa acgcgatatc gtaccccgag taagccgcac
     1321 ccgaggcttc gactgttacg ccgaatggag gattggtaag aataaaatcg aagtagccgt
     1381 agcgacacca gggtttagaa tggcgtttgc ccggttcgac aagcgagtcc atcagggcgc
     1441 gattaaacgt taagccaggg aagtgaccaa ggttaagttc tccaattaaa agcatacgag
     1501 ggttcttatc cgccccccag acactgatct tctgcgcatt ttccttacgc cacttcttga
     1561 tcacttcaat taaaaaggtc cccgatccgc acgcaggatc caggcattta gcagagctgg
     1621 gcgggtctac aaacgagact acctcacgta cgacttcatc aggagtcaga aagatcccaa
     1681 gtcccttgcg tacttctggt gtcaaaaact cacgaagagc cgccgaacga atatcataat
     1741 cgatctgtgt gaaattaatc ccgctgaaca gagcgttcaa ttgttctaag cactggtctg
     1801 aaaggtgaaa gtcacgatcg cgccataaac tcgtgctcca actactgtgg cgggacaggt
     1861 ggaatgcaaa gcgtttgcga atcgcgcttc ccgttgtagg ggcgatacca tcttcctcgg
     1921 acatttgctt taagaataag aacttcagca gttcctcaaa cgcctcttgc ggttgaagtc
     1981 catctaaatt gcgcatggta ttgtgggcac gacgataagt actagcaaga tcacccatgg
     2041 tatatctcct tcttaaagtt aaacaaaatt atttctagag gggaattgtt atccgctcac
     2101 aattccccta tagtgagtcg tattaatttc gcgggatcga gatctcgatc ctctacgccg
     2161 gacgcatcgt ggccggcatc accggcgcca caggtgcggt tgctggcgcc tatatcgccg
     2221 acatcaccga tggggaagat cgggctcgcc acttcgggct catgagcgct tgtttcggcg
     2281 tgggtatggt ggcaggcccc gtggccgggg gactgttggg cgccatctcc ttgcatgcac
     2341 cattccttgc ggcggcggtg ctcaacggcc tcaacctact actgggctgc ttcctaatgc
     2401 aggagtcgca taagggagag cgtcgagatc ccggacacca tcgaatggcg caaaaccttt
     2461 cgcggtatgg catgatagcg cccggaagag agtcaattca gggtggtgaa tgtgaaacca
     2521 gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg
     2581 gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg
     2641 gagctgaatt acattcccaa ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg
     2701 attggcgttg ccacctccag tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt
     2761 aaatctcgcg ccgatcaact gggtgccagc gtggtggtgt cgatggtaga acgaagcggc
     2821 gtcgaagcct gtaaagcggc ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc
     2881 attaactatc cgctggatga ccaggatgcc attgctgtgg aagctgcctg cactaatgtt
     2941 ccggcgttat ttcttgatgt ctctgaccag acacccatca acagtattat tttctcccat
     3001 gaagacggta cgcgactggg cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg
     3061 ctgttagcgg gcccattaag ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa
     3121 tatctcactc gcaatcaaat tcagccgata gcggaacggg aaggcgactg gagtgccatg
     3181 tccggttttc aacaaaccat gcaaatgctg aatgagggca tcgttcccac tgcgatgctg
     3241 gttgccaacg atcagatggc gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc
     3301 gttggtgcgg atatctcggt agtgggatac gacgataccg aagacagctc atgttatatc
     3361 ccgccgttaa ccaccatcaa acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc
     3421 ttgctgcaac tctctcaggg ccaggcggtg aagggcaatc agctgttgcc cgtctcactg
     3481 gtgaaaagaa aaaccaccct ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     3541 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     3601 cgcaattaat gtaagttagc tcactcatta ggcaccggga tctcgaccga tgcccttgag
     3661 agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact
     3721 tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tctgggtcat
     3781 tttcggcgag gaccgctttc gctggagcgc gacgatgatc ggcctgtcgc ttgcggtatt
     3841 cggaatcttg cacgccctcg ctcaagcctt cgtcactggt cccgccacca aacgtttcgg
     3901 cgagaagcag gccattatcg ccggcatggc ggccccacgg gtgcgcatga tcgtgctcct
     3961 gtcgttgagg acccggctag gctggcgggg ttgccttact ggttagcaga atgaatcacc
     4021 gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac
     4081 atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa acgcggaagt cagcgccctg
     4141 caccattatg ttccggatct gcatcgcagg atgctgctgg ctaccctgtg gaacacctac
     4201 atctgtatta acgaagcgct ggcattgacc ctgagtgatt tttctctggt cccgccgcat
     4261 ccataccgcc agttgtttac cctcacaacg ttccagtaac cgggcatgtt catcatcagt
     4321 aacccgtatc gtgagcatcc tctctcgttt catcggtatc attaccccca tgaacagaaa
     4381 tcccccttac acggaggcat cagtgaccaa acaggaaaaa accgccctta acatggcccg
     4441 ctttatcaga agccagacat taacgcttct ggagaaactc aacgagctgg acgcggatga
     4501 acaggcagac atctgtgaat cgcttcacga ccacgctgat gagctttacc gcagctgcct
     4561 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     4621 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     4681 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg
     4741 cttaactatg cggcatcaga gcagattgta ctgagagtgc accattgcgg tgtgaaatac
     4801 cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc tcgctcactg
     4861 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     4921 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     4981 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     5041 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     5101 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     5161 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     5221 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     5281 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     5341 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     5401 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     5461 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     5521 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     5581 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     5641 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgaacaat aaaactgtct
     5701 gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcttgc
     5761 tctaggccgc gattaaattc caacatggat gctgatttat atgggtataa atgggctcgc
     5821 gataatgtcg ggcaatcagg tgcgacaatc tatcgattgt atgggaagcc cgatgcgcca
     5881 gagttgtttc tgaaacatgg caaaggtagc gttgccaatg atgttacaga tgagatggtc
     5941 agactaaact ggctgacgga atttatgcct cttccgacca tcaagcattt tatccgtact
     6001 cctgatgatg catggttact caccactgcg atccccggga aaacagcatt ccaggtatta
     6061 gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc tggcagtgtt cctgcgccgg
     6121 ttgcattcga ttcctgtttg taattgtcct tttaacagcg atcgcgtatt tcgtctcgct
     6181 caggcgcaat cacgaatgaa taacggtttg gttgatgcga gtgattttga tgacgagcgt
     6241 aatggctggc ctgttgaaca agtctggaaa gaaatgcata aacttttgcc attctcaccg
     6301 gattcagtcg tcactcatgg tgatttctca cttgataacc ttatttttga cgaggggaaa
     6361 ttaataggtt gtattgatgt tggacgagtc ggaatcgcag accgatacca ggatcttgcc
     6421 atcctatgga actgcctcgg tgagttttct ccttcattac agaaacggct ttttcaaaaa
     6481 tatggtattg ataatcctga tatgaataaa ttgcagtttc atttgatgct cgatgagttt
     6541 ttctaagaat taattcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6601 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     6661 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     6721 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     6781 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     6841 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     6901 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     6961 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     7021 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     7081 ctacagggcg cgtcccattc gcca
//
