LOCUS       sequence-309624-        7736 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid for naturally occurring 6xMCE protein (without TM
            region).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7736)
  AUTHORS   Isom GL, Coudray N, MacRae MR, McManus CT, Ekiert DC, Bhabha G
  TITLE     LetB Structure Reveals a Tunnel for Lipid Transport across the 
            Bacterial Envelope.
  JOURNAL   Cell. 2020 Apr 30;181(3):653-664.e19. doi: 
            10.1016/j.cell.2020.03.030.
  PUBMED    32359438
REFERENCE   2  (bases 1 to 7736)
  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"; doi: 
            "10.1016/j.cell.2020.03"; journalName: "Cell"; date: "2020-04-30- 
            30"; volume: "181"; issue: "3"; pages: "653-664.e19"
COMMENT     Sequence Label: pBEL1753
FEATURES             Location/Qualifiers
     source          1..7736
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     331..350
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(370..447)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     653..672
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     756..775
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        756..774
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    775..799
                     /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             3049..3066
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3132..3150)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3146..3193
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3230..3685
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3317..3336)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3527..3548
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(3760..3793)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        3872..3976
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3977..4045,4046..4837)
                     /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(4195..4214)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5008..5596
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5497..5516
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5750..5767
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5782..5921
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5847..5866)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6007..6029
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6023..6214)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(7023..7736,1..369))
                     /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 gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc gccgctttac aggcttcgac
       61 gccgcttcgt tctaccatcg acaccaccac gctggcaccc agttgatcgg cgcgagattt
      121 aatcgccgcg acaatttgcg acggcgcgtg cagggccaga ctggaggtgg caacgccaat
      181 cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg ttgggaatgt aattcagctc
      241 cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa acgtggctgg cctggttcac
      301 cacgcgggaa acggtctgat aagagacacc ggcatactct gcgacatcgt ataacgttac
      361 tggtttcaca ttcaccaccc tgaattgact ctcttccggg cgctatcatg ccataccgcg
      421 aaaggttttg cgccattcga tggtgtccgg gatctcgacg ctctccctta tgcgactcct
      481 gcattaggaa gcagcccagt agtaggttga ggccgttgag caccgccgcc gcaaggaatg
      541 gtgcatgcaa ggagatggcg cccaacagtc ccccggccac ggggcctgcc accataccca
      601 cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg atcttcccca tcggtgatgt
      661 cggcgatata ggcgccagca accgcacctg tggcgccggt gatgccggcc acgatgcgtc
      721 cggcgtagag gatcgagatc tcgatcccgc gaaattaata cgactcacta taggggaatt
      781 gtgagcggat aacaattccc ctctagaaat aattttgttt aactttggcc caggcggcca
      841 ggagggtctc acatgggcgt ggacatccag gttcgctttg aatcaggaga tggtattcag
      901 gcaaataaga ccgaagtgtt gtacaaagga atttctgttg gtaaggtgac cgatttacac
      961 gtatcgaaag atatcaaggg tgtcgtcgcg acgatcgaga tcaagaaaga agcccaggaa
     1021 tacctttcga aggatactcg tttctggtta gttaagccgc gtgtctcact tgccggagtt
     1081 acaggcttag agacgttggt gtcaggggtt tacatcgcgg tggatcccgt gaaaggggag
     1141 aaggaggaac gctactttac tgcgttaaaa gaaccgccac ccttatctga caaattacca
     1201 ggtttacacc ttactttgaa ggctgatcgt ttaggaagtt tggagcaggg gagcccggta
     1261 ttttatcgtc aaattcaagt gggccaagtt aaatctttcc agttgggtga cgatcagcgt
     1321 actatcgaga tcaaggtaca tatcgaaccg gcttatgcgg atttagttcg taaacacacc
     1381 cgtttctgga acgcctctgg aatttcgatt agtgggggat tatcgggatt taaagtgcat
     1441 agcgaatccc ttttgacatt ggttgctggc gggatcgcgt tttcgacccc agaaaatcgt
     1501 accgattctc cgcctacaga ccccagcaag ccctttcgct tgtacgacga ctatgatgcc
     1561 gcgcaggccg gcttacgcgt caaacttaaa atgaacgacg tgagcgggat tgaccctggg
     1621 cgcactccgg taatgtttaa cggagttcaa gtcggtctgg tgaagtccat cgacatgggt
     1681 aaagactact cctccgctac tgctgacctg gcgatggacc cacgcgtcga agacatgctt
     1741 ttggaaggga ccgagttctg gacggtcaaa ccgagtatca gcttggccgg gattacaggt
     1801 cttgaggcgt tagtcaaggg aaactacatt gacgtgcgtt tcgcaaagtc tggggcgccg
     1861 tcccgcgaat ttacaatccg tccaaaagct cccccattga acaccgatgc gcctggactt
     1921 catttggtct taacgagcga caagttagga tctattgata tcggtgcacc tatcttgtac
     1981 cgtcaagttc gcgttggcag tgtccaaagt taccagctgt cgcgtgatcg tcaacgtgtc
     2041 gtggtaggtg ttcatatcga gcccgagtac gctcatcttg tcaacacgtc gacgcgcttt
     2101 tggaacagct ctgggattac acttacaggc aatttaagcg gggtacaagt caaatccgag
     2161 tcattgcaaa cgcttattac gggtggcatc tcctttgaca cgctggaccc caaggcccca
     2221 acagtcacca aggtccgccg cttcacactg ttcgatagtg aagaggctgc catggctcgc
     2281 ggggttgaga ttcagttgtc aatcgacaat gcggatggac tgcgtgaggg aacacccatt
     2341 cgctttaagg ggcttgacat cggtaaaatt gaatcggtcg agcttaaccc cgacttgagc
     2401 ggggtcttaa tgaaggcacg tttaacttcg gccggagagc gtgttgcccg ttccggtact
     2461 cgtttctggg ttgttcgtcc tgctttaggt cttttgcgca cagaaaacct gggaacactg
     2521 gtgtcagggc cgtacatcga agcgttacct agtagcactc ccggcgagcg ccaagcacgc
     2581 tttcaaacat tagccgaggc cccgaacctg ctgggccgtg aaaacggact gcgcttaact
     2641 ttatcagccc cccgtaaggg ctcaatcaaa cccggcaacc ttgtaacgta tcgccaaatt
     2701 cctgttggga aagtggtcga tttagcattg ggggaacagg ctgaccgtgt tctgatctct
     2761 atcctgattg agccgcgcta cgtgcctctt gtgcgtactg gctcgcgttt ttggaatgcg
     2821 agtgggttcg gcgtcgacgc atcgctgttc aaaggtttgt ctcttcgcac cgaatcgatg
     2881 gaggcgctga tggaaggcgg aattgcattc gcaacgccta ataacgcaca gatgggggaa
     2941 cctgcaaagc ctgggcaaac cttcgccctt ttcgacagtg ccaacgacga gtggttggag
     3001 tgggccccaa aaattcccct gaaggaaacc gcccgccgcg gcgagaccca tcaccaccat
     3061 caccattaag gccaggccgg ccgatccggc tgctaacaaa gcccgaaagg aagctgagtt
     3121 ggctgctgcc accgctgagc aataactagc ataacccctt ggggcctcta aacgggtctt
     3181 gaggggtttt ttgctgaaag gaggaactat atccggattg gcgaatggga cgcgccctgt
     3241 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3301 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3361 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3421 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     3481 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3541 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3601 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     3661 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaacgacg acctgcagcc
     3721 aagctagctt ggctggacgt aaactcctct tcagacctaa taacttcgta tagcatacat
     3781 tatacgaagt tatattaagg gttattgaat atgatcggaa ttgggctgca ggaattcgat
     3841 agcttggctg caggtcgacc tcgaggatgt gcgcggaacc cctatttgtt tatttttcta
     3901 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     3961 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     4021 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     4081 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     4141 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     4201 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     4261 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     4321 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     4381 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     4441 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     4501 gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat taactggcga
     4561 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     4621 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     4681 cggtgagcgt ggctctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     4741 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     4801 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     4861 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     4921 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     4981 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5041 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5101 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     5161 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     5221 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     5281 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     5341 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     5401 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     5461 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     5521 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     5581 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     5641 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     5701 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     5761 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     5821 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     5881 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     5941 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     6001 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     6061 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     6121 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     6181 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     6241 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     6301 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     6361 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     6421 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     6481 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     6541 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     6601 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     6661 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     6721 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     6781 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     6841 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     6901 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     6961 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     7021 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     7081 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     7141 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     7201 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     7261 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     7321 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     7381 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     7441 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     7501 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     7561 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     7621 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     7681 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaat
//
