LOCUS       sequence-398035-        5518 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of N-terminally His6-mCherry-tagged WzzE from 
            Pectobacterium atrosepticum in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5518)
  AUTHORS   Weckener M, Woodward LS, Clarke BR, Liu H, Ward PN, Le Bas A, Bhella
            D, Whitfield C, Naismith JH
  TITLE     The lipid linked oligosaccharide polymerase Wzy and its regulating 
            co-polymerase, Wzz, from enterobacterial common antigen biosynthesis
            form a complex.
  JOURNAL   Open Biol. 2023 Mar;13(3):220373. doi: 10.1098/rsob.220373. Epub 
            2023 Mar 22.
  PUBMED    36944376
REFERENCE   2  (bases 1 to 5518)
  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: "Open 
            Biol."; date: "2023-03"; volume: "13(3):220373. doi"; pages: " 
            10.1098/rsob.220373. Epub 2023 Mar 22"
COMMENT     Sequence Label: pHis-mCherry-WzzE
FEATURES             Location/Qualifiers
     source          1..5518
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     410..429
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(449..526)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     572..591
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        572..590
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    591..615
                     /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             671..688
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(843..861)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1111..1130
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2462..2506
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2544..2562)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2558..2605
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(2826..3485)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3400..3419
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3486..3588)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(3980..3997)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4114..4659)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(4194..4213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4884..5518,1..448))
                     /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"
ORIGIN
        1 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
       61 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
      121 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
      181 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
      241 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
      301 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
      361 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
      421 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
      481 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
      541 tctcccttat gcgactcctg cattaggaaa ttaatacgac tcactatagg ggaattgtga
      601 gcggataaca attcccctgt agaaataatt ttgtttaact ttaataagga gatataccat
      661 gggcagcagc catcaccatc atcaccacag ccaggatccg gtgagcaagg gcgaggagga
      721 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
      781 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
      841 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
      901 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
      961 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     1021 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     1081 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ctatgggctg
     1141 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     1201 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     1261 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     1321 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     1381 cggcggcatg aagcttgcca tggtgaaatc agagaacttg tctaccggga atgcgttgat
     1441 tgataacgaa ctggatatcc gtggtttatt ccgcctgctg tggcgtggaa aggtgtggcc
     1501 gatagcaatc ggcctgctgt ttgccgtcgt ggcattaggc tattcctatc ttgttaaaca
     1561 ggaatggggt gcaacatcga ttaccgacaa gccgacggtt aatatgttgg gcggctatta
     1621 ctcgcagcag caattcctac gtaacctcga cgctcgatcg ttttcctctc ctcagccaga
     1681 acaaccgtct atttcggctg gtgcttatga tgaattcatc atgcagttag cggcttacga
     1741 tacccgccgc gatttctggt tgcagactga ctattataag caacgtctgg aaggtgatga
     1801 aaaggctgat gcagccttgc tggatgagct ggtcaacaac attcagttta cggcgcgtga
     1861 tgacgggaaa aaaaccaatg actccgtgaa gctggtggcg gaaacctcct cagattccaa
     1921 cacgttgctt cgtcagtatg tggtttttgc cagtcagcgt gcagccaacc acttgaatga
     1981 agaaatcaag ggagcctggg ctgccagaac cattttcatg aagtcgcaga ttaagcgtca
     2041 ggaagcggta gcaaaagcga tttacgatcg tgaagtccgc agcgttgaac tggcgctgaa
     2101 aatcgcacaa cagcaaggta ttagccgtag ccagacggat acgccagcag atgaaatccc
     2161 cgcatcggaa atgttcctgc tcggcaggcc gatgcttcaa gcccgactgg aaacgttgca
     2221 gaccagcggc ccgcattatg aactggatta cgatcaaaat cgcgcgatgt tggcgacgct
     2281 gaacgttggc ccaacgcttg aggccagttt ccagacgtat cgttatttgc gtacaccgga
     2341 agaaccggtg aaacgcgaca gcccgcgccg ggcattcctg atggtgatgt ggggtgcgat
     2401 tggtgtgcta gtgggagcgg gtgttgccct ggctcgccgt ctgcgttagc tcgagtctgg
     2461 taaagaaacc gctgctgcga aatttgaacg ccagcacatg gactcgtcta ctagcgcagc
     2521 ttaattaacc taggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc
     2581 taaacgggtc ttgaggggtt ttttgctgaa acctcaggca tttgagaagc acacggtcac
     2641 actgcttccg gtagtcaata aaccggtaaa ccagcaatag acataagcgg ctatttaacg
     2701 accctgccct gaaccgacga ccgggtcgaa tttgctttcg aatttctgcc attcatccgc
     2761 ttattatcac ttattcaggc gtagcaacca ggcgtttaag ggcaccaata actgccttaa
     2821 aaaaattacg ccccgccctg ccactcatcg cagtactgtt gtaattcatt aagcattctg
     2881 ccgacatgga agccatcaca aacggcatga tgaacctgaa tcgccagcgg catcagcacc
     2941 ttgtcgcctt gcgtataata tttgcccata gtgaaaacgg gggcgaagaa gttgtccata
     3001 ttggccacgt ttaaatcaaa actggtgaaa ctcacccagg gattggctga gacgaaaaac
     3061 atattctcaa taaacccttt agggaaatag gccaggtttt caccgtaaca cgccacatct
     3121 tgcgaatata tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa
     3181 aacgtttcag tttgctcatg gaaaacggtg taacaagggt gaacactatc ccatatcacc
     3241 agctcaccgt ctttcattgc catacggaac tccggatgag cattcatcag gcgggcaaga
     3301 atgtgaataa aggccggata aaacttgtgc ttatttttct ttacggtctt taaaaaggcc
     3361 gtaatatcca gctgaacggt ctggttatag gtacattgag caactgactg aaatgcctca
     3421 aaatgttctt tacgatgcca ttgggatata tcaacggtgg tatatccagt gatttttttc
     3481 tccattttag cttccttagc tcctgaaaat ctcgataact caaaaaatac gcccggtagt
     3541 gatcttattt cattatggtg aaagttggaa cctcttacgt gccgatcaac gtctcatttt
     3601 cgccaaaagt tggcccaggg cttcccggta tcaacaggga caccaggatt tatttattct
     3661 gcgaagtgat cttccgtcac aggtatttat tcggcgcaaa gtgcgtcggg tgatgctgcc
     3721 aacttactga tttagtgtat gatggtgttt ttgaggtgct ccagtggctt ctgtttctat
     3781 cagctgtccc tcctgttcag ctactgacgg ggtggtgcgt aacggcaaaa gcaccgccgg
     3841 acatcagcgc tagcggagtg tatactggct tactatgttg gcactgatga gggtgtcagt
     3901 gaagtgcttc atgtggcagg agaaaaaagg ctgcaccggt gcgtcagcag aatatgtgat
     3961 acaggatata ttccgcttcc tcgctcactg actcgctacg ctcggtcgtt cgactgcggc
     4021 gagcggaaat ggcttacgaa cggggcggag atttcctgga agatgccagg aagatactta
     4081 acagggaagt gagagggccg cggcaaagcc gtttttccat aggctccgcc cccctgacaa
     4141 gcatcacgaa atctgacgct caaatcagtg gtggcgaaac ccgacaggac tataaagata
     4201 ccaggcgttt ccccctggcg gctccctcgt gcgctctcct gttcctgcct ttcggtttac
     4261 cggtgtcatt ccgctgttat ggccgcgttt gtctcattcc acgcctgaca ctcagttccg
     4321 ggtaggcagt tcgctccaag ctggactgta tgcacgaacc ccccgttcag tccgaccgct
     4381 gcgccttatc cggtaactat cgtcttgagt ccaacccgga aagacatgca aaagcaccac
     4441 tggcagcagc cactggtaat tgatttagag gagttagtct tgaagtcatg cgccggttaa
     4501 ggctaaactg aaaggacaag ttttggtgac tgcgctcctc caagccagtt acctcggttc
     4561 aaagagttgg tagctcagag aaccttcgaa aaaccgccct gcaaggcggt tttttcgttt
     4621 tcagagcaag agattacgcg cagaccaaaa cgatctcaag aagatcatct tattaatcag
     4681 ataaaatatt tctagatttc agtgcaattt atctcttcaa atgtagcacc tgaagtcagc
     4741 cccatacgat ataagttgta attctcatgt tagtcatgcc ccgcgcccac cggaaggagc
     4801 tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct
     4861 aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     4921 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg
     4981 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
     5041 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
     5101 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
     5161 gagatgtccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
     5221 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
     5281 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
     5341 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
     5401 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
     5461 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagaga
//
