LOCUS       sequence-309746-        8039 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expression plasmid for LetB PLL6 deletion (without TM region).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8039)
  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 8039)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pBEL1998
FEATURES             Location/Qualifiers
     source          1..8039
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     973..992
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1012..1089)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1295..1314
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1398..1417
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1398..1416
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1417..1441
                     /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             3994..4011
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4077..4095)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4091..4138
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4175..4630
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4262..4281)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4472..4493
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(4705..4738)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        4817..4921
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4922..4990,4991..5782)
                     /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(5140..5159)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5953..6541
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6442..6461
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6695..6712
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6727..6866
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6792..6811)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6952..6974
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6968..7159)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(7968..8039,1..1011))
                     /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 ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag
       61 ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg
      121 ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc
      181 ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt
      241 aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac
      301 gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc
      361 ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag
      421 acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc
      481 caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact
      541 gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc
      601 ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg
      661 ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat
      721 cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg
      781 cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc
      841 cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac
      901 tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg
      961 ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac
     1021 cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc
     1081 gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca
     1141 gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg
     1201 cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca
     1261 tgagcccgaa gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag
     1321 caaccgcacc tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga
     1381 tctcgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc
     1441 ccctctagaa ataattttgt ttaactttgg cccaggcggc caggagggtc tcacatgcag
     1501 gaccggggta ataccgtcac catcgacttt atgtcggcgg atggtattgt tccaggccgt
     1561 acgcctgttc gttatcaggg cgttgaagtc ggaacagtgc aggacatcag cctcagcgac
     1621 gatcttcgta agattgaagt caaggtcagc atcaagtccg atatgaaaga tgcgctgcgc
     1681 gaagagactc agttctggct ggtgacgcca aaagcatcgt tggcaggtgt ctccgggctg
     1741 gacgccctcg tcggtgggaa ctatatcggc atgatgcccg ggaaaggtaa agagcaggat
     1801 cactttgtcg cactcgatac ccaaccgaaa tatcggctgg acaatggcga tctgatgatc
     1861 cacctgcaag cccccgatct cggttcgctg aacagcggtt cattggtcta tttccgcaag
     1921 atcccggtgg gaaaagtcta cgactatgcc atcaatccca acaagcaagg cgtggtgatt
     1981 gatgtcctga tcgagcggcg ttttaccgat ctggtgaaaa aaggtagccg tttctggaac
     2041 gtttccggcg ttgatgccaa cgtcagtatc agtggcgcga aggtgaaact ggaaagcttg
     2101 gcggcactgg ttaacggtgc gattgccttc gattcaccag aagagtcgaa acctgccgag
     2161 gcggaagata cctttggtct gtatgaagat ctggcccaca gccagcgtgg cgtaataata
     2221 aaactggaac tgccgagtgg ggccggatta accgccgact cgacgccgtt aatgtatcag
     2281 gggctggaag tcggacagct gactaaactg gatttaaatc ctggtggtaa agtcacaggg
     2341 gaaatgaccg ttgatcccag cgtcgttacc ctgcttcgtg aaaatacccg catcgaatta
     2401 cgcaacccga aattatccct tagcgatgct aatctcagcg ccctgctgac cggcaaaacc
     2461 ttcgagctgg tgcccggcga tggcgagcca cgcaaagaat tcgttgttgt gccaggcgaa
     2521 aaagcactgc tgcatgaacc tgatgttctg acgctgaccc tgaccgcgcc ggaaagttac
     2581 ggtattgatg cgggtcagcc gctcattctt cacggcgtgc aggtaggcca ggttatcgat
     2641 cgtaaactca ccagcaaagg cgtcaccttt accgtcgcca tcgagcctca gcatcgcgaa
     2701 ctggtaaaag gcgatagcaa atttgtcgtc aacagccgtg tcgatgtgaa ggtggggctg
     2761 gatggcgttg agtttctcgg tgccagcgcc tcagaatgga tcaatggcgg gatccgtatt
     2821 ctgccgggcg ataaaggtga gatgaaagcc agctatccac tgtatgccaa tttggaaaaa
     2881 gcgctggaga acagccttag cgatttaccc accacaaccg tgagtttgag tgcagagacg
     2941 ctgccggatg tgcaggcagg atcggtagtg ctgtaccgta aatttgaagt tggtgaagtg
     3001 attaccgtgc gtccgcgagc taacgcgttt gacatcgatc tgcatattaa gccggagtat
     3061 cgcaaccttc tgaccagcaa tagcgtgttc tgggcagaag gcggggcgaa agttcagctg
     3121 aatggtagtg gtctgaccgt acaggcatcc ccgctctcca gagctctaaa gggagccatt
     3181 agcttcgaca acctcagcgg tgccagcgcc agtcagcgta aaggcgataa gcgtattctg
     3241 tatgcttccg aaacagcggc ccgtgcggtt ggcgggcaga ttacgcttca cgctttcgat
     3301 gccggaaaac tggcggtcgg gatgccaatt cgctatctcg gtattgacat cgggcaaatc
     3361 cagacgctgg atctgattac cgcacgcaat gaagtacagg caaaggcggt actttacccg
     3421 gaatatgtcc agacctttgc tcgcggtggt acgcgcttct cagtggtcac accgggaggg
     3481 catcttgata ctatcctcca gccgtatatc aacgtcgaac caggccgggg caatcctcgc
     3541 cgcgactttg aattacaaga ggccaccatt actgattcgc ggtacctgga tggcttaagc
     3601 attattgttg aagcgccgga agccggttcg ttaggcatcg gtacgcctgt gctgttccgt
     3661 ggtctggaag tcggtacggt tacaggaatg acgctgggga cattgtcaga tcgcgtgatg
     3721 attgcgatgc gcatcagtaa acgctatcaa cacctggtgc gtaacaattc cgtcttctgg
     3781 ttggcatcgg gttacagtct ggactttggt ctgacgggcg gcgtagtgaa aaccggcacc
     3841 tttaaccagt ttatccgtgg cggcatcgcc ttcgccacgc ctccgggtac gccactggca
     3901 ccgaaagccc aggaaggcaa acacttcctg ttgcaggaaa gtgaaccgaa agagtggcgt
     3961 gaatggggaa ctgcgcttcc caaaggcgag acccatcacc accatcacca ttaaggccag
     4021 gccggccgat ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc
     4081 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     4141 gaaaggagga actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc
     4201 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     4261 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     4321 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     4381 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     4441 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     4501 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     4561 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     4621 cttacaattt aggtggcact tttcggggaa cgacgacctg cagccaagct agcttggctg
     4681 gacgtaaact cctcttcaga cctaataact tcgtatagca tacattatac gaagttatat
     4741 taagggttat tgaatatgat cggaattggg ctgcaggaat tcgatagctt ggctgcaggt
     4801 cgacctcgag gatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4861 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     4921 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     4981 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5041 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5101 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5161 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5221 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5281 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5341 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5401 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5461 gcctgcagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     5521 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5581 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtggctc
     5641 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5701 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5761 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     5821 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     5881 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     5941 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6001 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6061 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     6121 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6181 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6241 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6301 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6361 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6421 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6481 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6541 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     6601 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     6661 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     6721 agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcaatgg
     6781 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac actccgctat
     6841 cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct gacgcgccct
     6901 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     6961 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     7021 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     7081 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     7141 ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc atgggggtaa
     7201 tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat gaacatgccc
     7261 ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg gaccagagaa
     7321 aaatcactca gggtcaatgc cagcgcttcg ttaatacaga tgtaggtgtt ccacagggta
     7381 gccagcagca tcctgcgatg cagatccgga acataatggt gcagggcgct gacttccgcg
     7441 tttccagact ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag
     7501 acgttttgca gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac
     7561 cagtaaggca accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca
     7621 cccgtggggc cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg
     7681 gaccagtgac gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc
     7741 cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg
     7801 gcacctgtcc tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca
     7861 tgccccgcgc ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag
     7921 atcccggtgc ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt
     7981 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggaga
//
