LOCUS       sequence-342007-        8024 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid for LetB PLL7 deletion (without TM region).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8024)
  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 8024)
  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: pBEL1999
FEATURES             Location/Qualifiers
     source          1..8024
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     970..989
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1009..1086)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1292..1311
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1395..1414
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1395..1413
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1414..1438
                     /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             3976..3993
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4059..4077)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4073..4120
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4157..4612
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4244..4263)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4454..4475
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(4687..4720)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4799..4903
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4904..4972,4973..5764)
                     /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(5122..5141)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5935..6523
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6424..6443
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6677..6694
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6709..6848
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6774..6793)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6934..6956
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6950..7141)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(7950..8024,1..1008))
                     /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 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
       61 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
      121 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
      181 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
      241 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
      301 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
      361 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
      421 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
      481 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
      541 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
      601 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
      661 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
      721 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
      781 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
      841 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
      901 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
      961 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     1021 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     1081 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     1141 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     1201 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     1261 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     1321 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
     1381 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
     1441 tctagaaata attttgttta actttggccc aggcggccag gagggtctca catgcaggac
     1501 cggggtaata ccgtcaccat cgactttatg tcggcggatg gtattgttcc aggccgtacg
     1561 cctgttcgtt atcagggcgt tgaagtcgga acagtgcagg acatcagcct cagcgacgat
     1621 cttcgtaaga ttgaagtcaa ggtcagcatc aagtccgata tgaaagatgc gctgcgcgaa
     1681 gagactcagt tctggctggt gacgccaaaa gcatcgttgg caggtgtctc cgggctggac
     1741 gccctcgtcg gtgggaacta tatcggcatg atgcccggga aaggtaaaga gcaggatcac
     1801 tttgtcgcac tcgataccca accgaaatat cggctggaca atggcgatct gatgatccac
     1861 ctgcaagccc ccgatctcgg ttcgctgaac agcggttcat tggtctattt ccgcaagatc
     1921 ccggtgggaa aagtctacga ctatgccatc aatcccaaca agcaaggcgt ggtgattgat
     1981 gtcctgatcg agcggcgttt taccgatctg gtgaaaaaag gtagccgttt ctggaacgtt
     2041 tccggcgttg atgccaacgt cagtatcagt ggcgcgaagg tgaaactgga aagcttggcg
     2101 gcactggtta acggtgcgat tgccttcgat tcaccagaag agtcgaaacc tgccgaggcg
     2161 gaagatacct ttggtctgta tgaagatctg gcccacagcc agcgtggcgt aataataaaa
     2221 ctggaactgc cgagtggggc cggattaacc gccgactcga cgccgttaat gtatcagggg
     2281 ctggaagtcg gacagctgac taaactggat ttaaatcctg gtggtaaagt cacaggggaa
     2341 atgaccgttg atcccagcgt cgttaccctg cttcgtgaaa atacccgcat cgaattacgc
     2401 aacccgaaat tatcccttag cgatgctaat ctcagcgccc tgctgaccgg caaaaccttc
     2461 gagctggtgc ccggcgatgg cgagccacgc aaagaattcg ttgttgtgcc aggcgaaaaa
     2521 gcactgctgc atgaacctga tgttctgacg ctgaccctga ccgcgccgga aagttacggt
     2581 attgatgcgg gtcagccgct cattcttcac ggcgtgcagg taggccaggt tatcgatcgt
     2641 aaactcacca gcaaaggcgt cacctttacc gtcgccatcg agcctcagca tcgcgaactg
     2701 gtaaaaggcg atagcaaatt tgtcgtcaac agccgtgtcg atgtgaaggt ggggctggat
     2761 ggcgttgagt ttctcggtgc cagcgcctca gaatggatca atggcgggat ccgtattctg
     2821 ccgggcgata aaggtgagat gaaagccagc tatccactgt atgccaattt ggaaaaagcg
     2881 ctggagaaca gccttagcga tttacccacc acaaccgtga gtttgagtgc agagacgctg
     2941 ccggatgtgc aggcaggatc ggtagtgctg taccgtaaat ttgaagttgg tgaagtgatt
     3001 accgtgcgtc cgcgagctaa cgcgtttgac atcgatctgc atattaagcc ggagtatcgc
     3061 aaccttctga ccagcaatag cgtgttctgg gcagaaggcg gggcgaaagt tcagctgaat
     3121 ggtagtggtc tgaccgtaca ggcatccccg ctctccagag ctctaaaggg agccattagc
     3181 ttcgacaacc tcagcggtgc cagcgccagt cagcgtaaag gcgataagcg tattctgtat
     3241 gcttccgaaa cagcggcccg tgcggttggc gggcagatta cgcttcacgc tttcgatgcc
     3301 ggaaaactgg cggtcgggat gccaattcgc tatctcggta ttgacatcgg gcaaatccag
     3361 acgctggatc tgattaccgc acgcaatgaa gtacaggcaa aggcggtact ttacccggaa
     3421 tatgtccaga cctttgctcg cggtggtacg cgcttctcag tggtcacacc gcaaatttcg
     3481 gcagctggcg ttgagcatct tgatactatc ctccagccgt atatcaacgt cgaaccaggc
     3541 cggggcaatc ctcgccgcga ctttgaatta caagaggcca ccattactga ttcgcggtac
     3601 ctggatggct taagcattat tgttgaagcg ccggaagccg gttcgttagg catcggtacg
     3661 cctgtgctgt tccgtggtct ggaagtcggt acggttacag gaatgacgct ggggacattg
     3721 tcagatcgcg tgatgattgc gatgcgcatc agtaaacgct atcaacacct ggtgcgtaac
     3781 aattccgtct tctggttggc atcgggttac ggtggcggca cctttaacca gtttatccgt
     3841 ggcggcatcg ccttcgccac gcctccgggt acgccactgg caccgaaagc ccaggaaggc
     3901 aaacacttcc tgttgcagga aagtgaaccg aaagagtggc gtgaatgggg aactgcgctt
     3961 cccaaaggcg agacccatca ccaccatcac cattaaggcc aggccggccg atccggctgc
     4021 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata
     4081 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc
     4141 cggattggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     4201 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     4261 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4321 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     4381 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     4441 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     4501 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     4561 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
     4621 cttttcgggg aacgacgacc tgcagccaag ctagcttggc tggacgtaaa ctcctcttca
     4681 gacctaataa cttcgtatag catacattat acgaagttat attaagggtt attgaatatg
     4741 atcggaattg ggctgcagga attcgatagc ttggctgcag gtcgacctcg aggatgtgcg
     4801 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     4861 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     4921 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     4981 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     5041 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     5101 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     5161 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     5221 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     5281 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     5341 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     5401 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     5461 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     5521 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     5581 ctggtttatt gctgataaat ctggagccgg tgagcgtggc tctcgcggta tcattgcagc
     5641 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     5701 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     5761 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     5821 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     5881 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     5941 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6001 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     6061 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     6121 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     6181 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     6241 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     6301 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     6361 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     6421 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     6481 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     6541 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     6601 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     6661 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     6721 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
     6781 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
     6841 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     6901 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     6961 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
     7021 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
     7081 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     7141 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     7201 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     7261 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     7321 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     7381 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     7441 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     7501 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     7561 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     7621 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     7681 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     7741 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     7801 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     7861 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     7921 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     7981 cgtgccagct gcattaatga atcggccaac gcgcggggag aggc
//
