LOCUS       sequence-356908-        7678 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid for LetB Delta-Ring 2 (without TM region).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7678)
  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 7678)
  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: pBEL1683
FEATURES             Location/Qualifiers
     source          1..7678
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..467
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(99..118)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     309..330
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(542..575)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        654..758
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(759..827,828..1619)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(977..996)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1790..2378
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2279..2298
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2532..2549
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2564..2703
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2629..2648)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2789..2811
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2805..2996)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3805..4887)
                     /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"
     primer_bind     4849..4868
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4888..4965)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5171..5190
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5274..5293
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5274..5292
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5293..5317
                     /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             7514..7531
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7592..7610)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      7606..7653
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
       61 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
      121 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
      181 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
      241 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
      301 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
      361 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
      421 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
      481 cggggaacga cgacctgcag ccaagctagc ttggctggac gtaaactcct cttcagacct
      541 aataacttcg tatagcatac attatacgaa gttatattaa gggttattga atatgatcgg
      601 aattgggctg caggaattcg atagcttggc tgcaggtcga cctcgaggat gtgcgcggaa
      661 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
      721 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
      781 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
      841 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
      901 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
      961 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     1021 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     1081 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     1141 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     1201 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     1261 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt
     1321 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     1381 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     1441 ttattgctga taaatctgga gccggtgagc gtggctctcg cggtatcatt gcagcactgg
     1501 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     1561 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     1621 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     1681 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     1741 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     1801 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     1861 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     1921 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     1981 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     2041 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     2101 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     2161 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     2221 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     2281 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     2341 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     2401 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     2461 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     2521 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc
     2581 tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc
     2641 tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct
     2701 gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     2761 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     2821 tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat
     2881 tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat
     2941 gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat
     3001 gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg
     3061 atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt
     3121 aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag
     3181 cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag
     3241 atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg
     3301 aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt
     3361 cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct
     3421 agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc catgccggcg
     3481 ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg
     3541 agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga
     3601 aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg
     3661 ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag
     3721 ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc
     3781 taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3841 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag
     3901 ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc
     3961 ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt
     4021 gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac
     4081 cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc
     4141 catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat
     4201 ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat
     4261 ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact
     4321 taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc
     4381 cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac
     4441 atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc
     4501 atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc
     4561 cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag
     4621 ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact
     4681 ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt
     4741 gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac
     4801 gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc
     4861 gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg
     4921 ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct
     4981 ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca
     5041 ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg
     5101 ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat
     5161 cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga
     5221 tgccggccac gatgcgtccg gcgtagagga tcgagatctc gatcccgcga aattaatacg
     5281 actcactata ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa
     5341 ctttggccca ggagatatac atatgcagga ccggggtaat accgtcacca tcgactttat
     5401 gtcggcggat ggtattgttc caggccgtac gcctgttcgt tatcagggcg ttgaagtcgg
     5461 aacagtgcag gatatcagcc tcagcgacga tcttcgtaag attgaagtca aggtcagcat
     5521 caagtccgat atgaaagatg cgctgcgcga agagactcag ttctggctgg tgacgccaaa
     5581 agcatcgttg gcaggtgtct ccgggctgga cgccctcgtc ggtgggaact atatcggcat
     5641 gatgccgggt aaaggtaaag agcaggatca ctttgtcgca ctcgataccc aaccgaaata
     5701 tcggctggac aatggcggcg taataataaa actggaactg ccgagtgggg ccggattaac
     5761 cgccgactcg acgccgttaa tgtatcaggg gctggaagtc ggacagctga ctaaactgga
     5821 tttaaatcct ggtggtaaag tcacagggga aatgaccgtt gatcccagcg tcgttaccct
     5881 gcttcgtgaa aatacccgca tcgaattacg caacccgaaa ttatccctta gcgatgctaa
     5941 tctcagcgcc ctgctgaccg gcaaaacctt cgagctggtg cccggcgatg gcgagccacg
     6001 caaagagttc gttgttgtgc caggcgaaaa agcactgctg catgaacctg atgttctgac
     6061 gctgaccctg accgcgccgg aaagttacgg tattgatgcg ggtcagccgc tcattcttca
     6121 cggcgtgcag gtaggccagg ttatcgatcg taaactcacc agcaaaggcg tcacctttac
     6181 cgtcgccatc gagcctcagc atcgcgaact ggtaaaaggc gatagcaaat ttgtcgtcaa
     6241 cagccgtgtc gatgtgaagg tggggctgga tggcgttgag tttctcggtg ccagcgcctc
     6301 agaatggatc aatggcggga tacgtattct gccgggcgat aaaggtgaga tgaaagccag
     6361 ctatccactg tatgccaatt tggaaaaagc gctggagaac agccttagcg atttacccac
     6421 cacaaccgtg agtttgagtg cagagacgct gccggatgtg caggcaggat cggtagtgct
     6481 gtaccgtaaa tttgaagttg gtgaagtgat taccgtgcgt ccgcgagcta acgcgtttga
     6541 tatcgatctg catattaagc cggagtatcg caaccttctg accagcaata gcgtgttctg
     6601 ggcagaaggc ggggcgaaag ttcagctgaa tggtagtggt ctgaccgtac aggcatcccc
     6661 gctctccaga gcattaaagg gagccattag cttcgacaac ctcagcggtg ccagcgccag
     6721 tcagcgtaaa ggcgataagc gtattctgta tgcttccgaa acagcggccc gtgcggttgg
     6781 cgggcagatt acgcttcacg ctttcgatgc cggaaaactg gcggtcggga tgccaattcg
     6841 ctatctcggt attgatatcg ggcaaatcca gacgctggat ctgattaccg cacgcaatga
     6901 agtacaggca aaggcggtac tttacccgga atatgtccag acctttgctc gcggtggtac
     6961 gcgcttctca gtggtcacac cgcaaatttc ggcagctggc gttgagcatc ttgatactat
     7021 cctccagccg tatatcaacg tcgaaccagg ccggggcaat cctcgccgcg actttgaatt
     7081 acaagaggcc accattactg attcgcgtta cctggatggc ttaagcatta ttgttgaagc
     7141 gccggaagcc ggttcgttag gcatcggtac gcctgtgctg ttccgtggtc tggaagtcgg
     7201 tacggttaca ggaatgacgc tggggacatt gtcagatcgc gtgatgattg cgatgcgcat
     7261 cagtaaacgc tatcaacacc tggtgcgtaa caattccgtg ttctggttgg catcgggtta
     7321 cagtctggac tttggtctga cgggcggcgt agtgaaaacc ggcaccttta accagtttat
     7381 ccgtggcggc atcgccttcg ccacgcctcc gggtacgcca ctggcaccga aagcccagga
     7441 aggcaaacac ttcctgttgc aggaaagtga accgaaagag tggcgtgaat ggggaactgc
     7501 gcttcccaaa ggccatcacc accatcacca ttaaggccgg ccgatccggc tgctaacaaa
     7561 gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
     7621 ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggat
//
