LOCUS       sequence-333949-        8658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expresson of LetA-[LetB PLL6->3] operon from E. coli. N-terminal
            his tag on LetA, araBAD promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8658)
  AUTHORS   Vieni C, Coudray N, Isom GL, Bhabha G, Ekiert DC
  TITLE     Role of Ring6 in the Function of the E. coli MCE Protein LetB.
  JOURNAL   J Mol Biol. 2022 Jan 22;434(7):167463. doi: 
            10.1016/j.jmb.2022.167463.
  PUBMED    35077766
REFERENCE   2  (bases 1 to 8658)
  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: "J Mol 
            Biol."; date: "2022-01-22"; volume: "434(7):167463. doi"; pages: " 
            10.1016/j.jmb.2022.167463"
COMMENT     Sequence Label: pBEL2197
FEATURES             Location/Qualifiers
     source          1..8658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(60..146)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     299..316
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(4307..4327)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             complement(4340..4363)
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     promoter        complement(4400..4684)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(4458..4477)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             4711..5589
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     primer_bind     complement(5769..5788)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(5926..5948)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6114..6131)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6303..6336
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(6433..7020)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6513..6532)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     rep_origin      complement(7131..7586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7268..7289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7480..7499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(join(7628..8419,8420..8488))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8251..8270
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8489..8580)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(8599..8626)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 cgtcctgccc gccaccctcc gggccgttgc ttcgcaacgt tcaaatccgc tcccggcgga
       61 tttgtcctac tcaggagagc gttcaccgac aaacaacaga taaaacgaaa ggcccagtct
      121 ttcgactgag cctttcgttt tatttgatgc ctggcagttc cctactctcg catggggaga
      181 ccccacacta ccatcggcgc tacggcgttt cacttctgag ttcggcatgg ggtcaggtgg
      241 gaccaccgcg ctactgccgc caggcaaatt ctgttttatc agaccgcttc tgcgttctga
      301 tttaatctgt atcaggctga aaatcttctc tcatccgcca aaacagccaa gcttacaacc
      361 attcctcaga ccaggaccaa aacgaaaaaa gacgcttttc agcgtctctt ttctggaatt
      421 tggtacttat ttgggaagcg cagttcccca ttcacgccac tctttcggtt cactttcctg
      481 caacaggaag tgtttgcctt cctgggcttt cggtgccagt ggcgtacccg gaggcgtggc
      541 gaaggcgatg ccgccacgga taaactggtt aaaggtgccg gttttcacta cgccgcccgt
      601 cagaccaaag tccagactgt aacccgatgc caaccagaag acggaattgt tacgcaccag
      661 gtgttgatag cgtttactga tgcgcatcgc aatcatcacg cgatctgaca atgtccccag
      721 cgtcattcct gtaaccgtac cgacttccag accacggaac agcacaggcg taccgatgcc
      781 taacgaaccg gcttccggcg cttcaacaat aatgcttaag ccatccaggt aacgcgaatc
      841 agtaatggtg gcctcttgta attcaaagtc gcggcgagga ttgccccggc ctggttcgac
      901 gttgatatac ggggtcagca gggcgctgag attagcatcg ctaagggata atttcggtgt
      961 gaccactgag aagcgcgtac caccgcgagc aaaggtctgg acatattccg ggtaaagtac
     1021 cgcctttgcc tgtacttcat tgcgtgcggt aatcagatcc agcgtctgga tttgcccgat
     1081 atcaataccg agatagcgaa ttggcatccc gaccgccagt tttccggcat cgaaagcgtg
     1141 aagcgtaatc tgcccgccaa ccgcacgggc cgctgtttcg gaagcataca gaatacgctt
     1201 atcgccttta cgctgactgg cgctggcacc gctgaggttg tcgaagctaa tggctccctt
     1261 taatgctctg gagagcgggg atgcctgtac ggtcagacca ctaccattca gctgaacttt
     1321 cgccccgcct tctgcccaga acacgctatt gctggtcaga aggttgcgat actccggctt
     1381 aatatgcaga tcgatatcaa acgcgttagc tcgcggacgc acggtaatca cttcaccaac
     1441 ttcaaattta cggtacagca ctaccgatcc tgcctgcaca tccggcagcg tctctgcact
     1501 caaactcacg gttgtggtgg gtaaatcgct aaggctgttc tccagcgctt tttccaaatt
     1561 ggcatacagt ggatagctgg ctttcatctc acctttatcg cccggcagaa tacgtatccc
     1621 gccattgatc cattctgagg cgctggcacc gagaaactca acgccatcca gccccacctt
     1681 cacatcgaca cggctgttga cgacaaattt gctatcgcct tttaccagtt cgcgatgctg
     1741 aggctcgatg gcgacggtaa aggtgacgcc tttgctggtg agtttacgat cgataacctg
     1801 gcctacctgc acgccgtgaa gaatgagcgg ctgacccgca tcaataccgt aactttccgg
     1861 cgcggtcagg gtcagcgtca gaacatcagg ttcatgcagc agtgcttttt cgcctggcac
     1921 aacaacgaac tctttgcgtg gctcgccatc gccgggcacc agctcgaagg ttttgccggt
     1981 cagcagggcg ctgagattag catcgctaag ggataatttc gggttgcgta attcgatgcg
     2041 ggtattttca cgaagcaggg taacgacgct gggatcaacg gtcatttccc ctgtgacttt
     2101 accaccagga tttaaatcca gtttagtcag ctgtccgact tccagcccct gatacattaa
     2161 cggcgtcgag tcggcggtta atccggcccc actcggcagt tccagtttta ttattacgcc
     2221 acgctggctg tgggccagat cttcatacag accaaaggta tcttccgcct cggcaggttt
     2281 cgactcttct ggtgaatcga aggcaatcgc accgttaacc agtgccgcca ggctttccag
     2341 tttcaccttc gcgccactga tactgacgtt ggcatcaacg ccggaaacgt tccagaaacg
     2401 gctacctttt ttcaccagat cggtaaaacg ccgctcgatc aggacatcaa tcaccacgcc
     2461 ttgcttgttg ggattgatgg catagtcgta gacttttccc accgggatct tgcggaaata
     2521 gaccaatgaa ccgctgttca gcgaaccgag atcgggggct tgcaggtgga tcatcagatc
     2581 gccattgtcc agccgatatt tcggttgggt atcgagtgcg acaaagtgat cctgctcttt
     2641 acctttaccc ggcatcatgc cgatatagtt cccaccgacg agggcgtcca gcccggagac
     2701 acctgccaac gatgcttttg gcgtcaccag ccagaactga gtctcttcgc gcagcgcatc
     2761 tttcatatcg gacttgatgc tgaccttgac ttcaatctta cgaagatcgt cgctgaggct
     2821 gatatcctgc actgttccga cttcaacgcc ctgataacga acaggcgtac ggcctggaac
     2881 aataccatcc gccgacataa agtcgatggt gacggtatta ccccggtcct gataactgtc
     2941 ccaaatcagc caactggcaa tcattagcgc gatgaacggc agcagccaga aaggtgagat
     3001 acggcgttta tttttaatct gcgcttcagt cgtcgaagcg ggcgtttcct gactcatgtg
     3061 catcccaaag taagcggctg tccagccatt ccacagcaag aatagtcaaa attaccgctg
     3121 cgccgaaata aaacgcagcc ggtcccatag taaaagcgag gatctgatcg cgattaatca
     3181 gcgacatggt taaagatatg acaaacaggt cgagcatcga ccagcgacca atccaggtca
     3241 ccatccgcag taacagaatg cgtgtgcgta aaccttgctg gcatttaaaa tgaatgctca
     3301 acagtaaagt gaacatgacg atcactttag taaacggtac cagaatactg gcgataaaca
     3361 cgattcctgc aaccgcaatg ttgctacttg ccagcgacat aattccggaa agaattgtat
     3421 cttcctgccg tcctccgttc agataaatga tagaaatagg caacaggttg gcaggtaaca
     3481 ataaaacgat tgacgctaac agcgccgccc agcatttttg cagactatga cggcgacgca
     3541 ggcgtagcgg gatatggcaa cgcgggcagc gaccacgctg atctggatag ccggtaaaat
     3601 ggcacccaag acagacacga agtttctcgt ccctacgcgt agcggggcgc tgcggataaa
     3661 atcgctccca cagttcttcg acattaagat gtgacaacgt caccgtcgtt aaaatcacca
     3721 acgccacaaa agagaacaag ccgacaccag cctggatatg ggcataatcc tgtaccttta
     3781 tagaagcaac gccaatgccg accaggtaga tatcgagcat tacccactct ttaagtcgct
     3841 caagcatcag cagtaccgga cgtaaattca ttcccagtcg gttaccaaac cataaataag
     3901 ctatggaggt caccagaatg agcggggcac caataacgca gaaaaagacc atcgaccccg
     3961 ttatcgcatc gccctgtttg gtcatttgcc agatgccttg cataacgttg gcgtcaatac
     4021 gaatgcctaa cagccagata tgcaacagcg gttcgcccca ggcaaacggc ataagcaaca
     4081 gcatggtgaa ggccattgcc gccaggcgcg ttagcgacca gtcgcgcccg tcacgaattt
     4141 ttgcctgaca gcgcggacaa taggcacttt gatgagaatt tatctcgggc aggctaaaca
     4201 gcatgtcaca ttgcgggcaa cgttggtaat caccacgcgg cagttcctcg ccgattgccc
     4261 tcactgttat cttttttgtc ggcgtaattt gtggtgtgtt aagagccatc tgaaaataca
     4321 gattttcgtg ttgatgctga tggtgatggt ggtgatggtg atgcataatt cacccttcgg
     4381 ctagcccaaa aaaacgggta tggagaaaca gtagagagtt gcgataaaaa gcgtcaggta
     4441 ggatccgcta atcttatgga taaaaatgct atggcatagc aaagtgtgac gccgtgcaaa
     4501 taatcaatgt ggacttttct gccgtgatta tagacacttt tgttacgcgt ttttgtcatg
     4561 gctttggtcc cgctttgtta cagaatgctt ttaataagcg gggttaccgg tttggttagc
     4621 gagaagagcc agtaaaagac gcagtgacgg caatgtctga tgcaatatgg acaattggtt
     4681 tcttctctga atggcgggag tatgaaaagt atggctgaag cgcaaaatga tcccctgctg
     4741 ccgggatact cgtttaatgc ccatctggtg gcgggtttaa cgccgattga ggccaacggt
     4801 tatctcgatt tttttatcga ccgaccgctg ggaatgaaag gttatattct caatctcacc
     4861 attcgcggtc agggggtggt gaaaaatcag ggacgagaat ttgtttgccg accgggtgat
     4921 attttgctgt tcccgccagg agagattcat cactacggtc gtcatccgga ggctcgcgaa
     4981 tggtatcacc agtgggttta ctttcgtccg cgcgcctact ggcatgaatg gcttaactgg
     5041 ccgtcaatat ttgccaatac ggggttcttt cgcccggatg aagcgcacca gccgcatttc
     5101 agcgacctgt ttgggcaaat cattaacgcc gggcaagggg aagggcgcta ttcggagctg
     5161 ctggcgataa atctgcttga gcaattgtta ctgcggcgca tggaagcgat taacgagtcg
     5221 ctccatccac cgatggataa tcgggtacgc gaggcttgtc agtacatcag cgatcacctg
     5281 gcagacagca attttgatat cgccagcgtc gcacagcatg tttgcttgtc gccgtcgcgt
     5341 ctgtcacatc ttttccgcca gcagttaggg attagcgtct taagctggcg cgaggaccaa
     5401 cgtatcagcc aggcgaagct gcttttgagc accacccgga tgcctatcgc caccgtcggt
     5461 cgcaatgttg gttttgacga tcaactctat ttctcgcggg tatttaaaaa atgcaccggg
     5521 gccagcccga gcgagttccg tgccggttgt gaagaaaaag tgaatgatgt agccgtcaag
     5581 ttgtcataat tggtaacgaa tcagacaatt gacggcttga cggagtagca tagggtttgc
     5641 agaatccctg cttcgtccat ttgacaggca cattatgcat cgatgataag ctgtcaaaca
     5701 tgagcagatc ctctacgccg gacgcatcgt ggccggcatc accggcgcca caggtgcggt
     5761 tgctggcgcc tatatcgccg acatcaccga tggggaagat cgggctcgcc acttcgggct
     5821 catgagcgct tgtttcggcg tgggtatggt ggcaggcccc gtggccgggg gactgttggg
     5881 cgccatctcc ttctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     5941 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     6001 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgaccctat gcggtgtgaa
     6061 ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc
     6121 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     6181 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtc ctcgaggtcg
     6241 acctgcagcc aagctatcga attcctgcag cccaattccg atcatattca ataaccctta
     6301 atataacttc gtataatgta tgctatacga agttattagg tctgaagagg agtttacgtc
     6361 cagccaagct aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     6421 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     6481 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     6541 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     6601 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     6661 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     6721 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     6781 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     6841 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     6901 gagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     6961 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     7021 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     7081 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattgtaaa
     7141 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     7201 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagcccgaga tagggttgag
     7261 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     7321 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccca aatcaagttt
     7381 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     7441 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     7501 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     7561 gcttaatgcg ccgctacagg gcgcgtaaat caatctaaag tatatatgag taaacttggt
     7621 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     7681 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     7741 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     7801 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     7861 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     7921 tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg
     7981 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     8041 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     8101 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     8161 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     8221 cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa
     8281 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     8341 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     8401 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     8461 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     8521 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     8581 agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt aatttgatgc
     8641 ctggcagttt atggcggg
//
