LOCUS       sequence-342010-        8012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression plasmid for LetB PLL1 deletion (without TM region).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8012)
  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 8012)
  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: pBEL1993
FEATURES             Location/Qualifiers
     source          1..8012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     304..323
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     557..574
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    589..728
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(654..673)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     814..836
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(830..1021)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1830..2912)
                     /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     2874..2893
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2913..2990)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     3196..3215
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     3299..3318
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3299..3317
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3318..3342
                     /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             5868..5885
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5951..5969)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5965..6012
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      6049..6504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6136..6155)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6346..6367
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(6579..6612)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6691..6795
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6796..6864,6865..7656)
                     /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(7014..7033)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(7827..8012,1..403)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
       61 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
      121 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
      181 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
      241 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
      301 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
      361 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
      421 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
      481 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
      541 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
      601 ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc
      661 tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca
      721 tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
      781 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
      841 caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa
      901 gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg
      961 ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca
     1021 ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag
     1081 agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg
     1141 agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat
     1201 gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga
     1261 tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa
     1321 cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt
     1381 cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc
     1441 agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc
     1501 cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt
     1561 gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc
     1621 agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt
     1681 gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga
     1741 aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt gcctaatgag
     1801 tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     1861 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     1921 gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc cttcaccgcc
     1981 tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag gcgaaaatcc
     2041 tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc gtcgtatccc
     2101 actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg cattgcgccc
     2161 agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc attcagcatt
     2221 tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc cgctatcggc
     2281 tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg cgccgagaca
     2341 gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac cagatgctcc
     2401 acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg tgtctggtca
     2461 gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc aatggcatcc
     2521 tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag aagattgtgc
     2581 accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac cacgctggca
     2641 cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc gtgcagggcc
     2701 agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg ttgtgccacg
     2761 cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg cgttttcgca
     2821 gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac accggcatac
     2881 tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg actctcttcc
     2941 gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc cgggatctcg
     3001 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     3061 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     3121 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     3181 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     3241 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta
     3301 atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg
     3361 tttaactttg gcccaggcgg ccaggagggt ctcacatgca ggaccggggt aataccgtca
     3421 ccatcgactt tatgtcggcg gatggtattg ttccaggccg tacgcctgtt cgttatcagg
     3481 gcgttgaagt cggaacagtg caggacatca gcctcagcga cgatcttcgt aagattgaag
     3541 tcaaggtcag catcaagtcc gatatgaaag atgcgctgcg cgaagagact cagttctggc
     3601 tggtgacggg agggaactat atcggcatga tgcccgggaa aggtaaagag caggatcact
     3661 ttgtcgcact cgatacccaa ccgaaatatc ggctggacaa tggcgatctg atgatccacc
     3721 tgcaagcccc cgatctcggt tcgctgaaca gcggttcatt ggtctatttc cgcaagatcc
     3781 cggtgggaaa agtctacgac tatgccatca atcccaacaa gcaaggcgtg gtgattgatg
     3841 tcctgatcga gcggcgtttt accgatctgg tgaaaaaagg tagccgtttc tggaacgttt
     3901 ccggcgttga tgccaacgtc agtatcagtg gcgcgaaggt gaaactggaa agcttggcgg
     3961 cactggttaa cggtgcgatt gccttcgatt caccagaaga gtcgaaacct gccgaggcgg
     4021 aagatacctt tggtctgtat gaagatctgg cccacagcca gcgtggcgta ataataaaac
     4081 tggaactgcc gagtggggcc ggattaaccg ccgactcgac gccgttaatg tatcaggggc
     4141 tggaagtcgg acagctgact aaactggatt taaatcctgg tggtaaagtc acaggggaaa
     4201 tgaccgttga tcccagcgtc gttaccctgc ttcgtgaaaa tacccgcatc gaattacgca
     4261 acccgaaatt atcccttagc gatgctaatc tcagcgccct gctgaccggc aaaaccttcg
     4321 agctggtgcc cggcgatggc gagccacgca aagaattcgt tgttgtgcca ggcgaaaaag
     4381 cactgctgca tgaacctgat gttctgacgc tgaccctgac cgcgccggaa agttacggta
     4441 ttgatgcggg tcagccgctc attcttcacg gcgtgcaggt aggccaggtt atcgatcgta
     4501 aactcaccag caaaggcgtc acctttaccg tcgccatcga gcctcagcat cgcgaactgg
     4561 taaaaggcga tagcaaattt gtcgtcaaca gccgtgtcga tgtgaaggtg gggctggatg
     4621 gcgttgagtt tctcggtgcc agcgcctcag aatggatcaa tggcgggatc cgtattctgc
     4681 cgggcgataa aggtgagatg aaagccagct atccactgta tgccaatttg gaaaaagcgc
     4741 tggagaacag ccttagcgat ttacccacca caaccgtgag tttgagtgca gagacgctgc
     4801 cggatgtgca ggcaggatcg gtagtgctgt accgtaaatt tgaagttggt gaagtgatta
     4861 ccgtgcgtcc gcgagctaac gcgtttgaca tcgatctgca tattaagccg gagtatcgca
     4921 accttctgac cagcaatagc gtgttctggg cagaaggcgg ggcgaaagtt cagctgaatg
     4981 gtagtggtct gaccgtacag gcatccccgc tctccagagc tctaaaggga gccattagct
     5041 tcgacaacct cagcggtgcc agcgccagtc agcgtaaagg cgataagcgt attctgtatg
     5101 cttccgaaac agcggcccgt gcggttggcg ggcagattac gcttcacgct ttcgatgccg
     5161 gaaaactggc ggtcgggatg ccaattcgct atctcggtat tgacatcggg caaatccaga
     5221 cgctggatct gattaccgca cgcaatgaag tacaggcaaa ggcggtactt tacccggaat
     5281 atgtccagac ctttgctcgc ggtggtacgc gcttctcagt ggtcacaccg caaatttcgg
     5341 cagctggcgt tgagcatctt gatactatcc tccagccgta tatcaacgtc gaaccaggcc
     5401 ggggcaatcc tcgccgcgac tttgaattac aagaggccac cattactgat tcgcggtacc
     5461 tggatggctt aagcattatt gttgaagcgc cggaagccgg ttcgttaggc atcggtacgc
     5521 ctgtgctgtt ccgtggtctg gaagtcggta cggttacagg aatgacgctg gggacattgt
     5581 cagatcgcgt gatgattgcg atgcgcatca gtaaacgcta tcaacacctg gtgcgtaaca
     5641 attccgtctt ctggttggca tcgggttaca gtctggactt tggtctgacg ggcggcgtag
     5701 tgaaaaccgg cacctttaac cagtttatcc gtggcggcat cgccttcgcc acgcctccgg
     5761 gtacgccact ggcaccgaaa gcccaggaag gcaaacactt cctgttgcag gaaagtgaac
     5821 cgaaagagtg gcgtgaatgg ggaactgcgc ttcccaaagg cgagacccat caccaccatc
     5881 accattaagg ccaggccggc cgatccggct gctaacaaag cccgaaagga agctgagttg
     5941 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     6001 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
     6061 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     6121 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     6181 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     6241 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     6301 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     6361 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     6421 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     6481 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaacgacga cctgcagcca
     6541 agctagcttg gctggacgta aactcctctt cagacctaat aacttcgtat agcatacatt
     6601 atacgaagtt atattaaggg ttattgaata tgatcggaat tgggctgcag gaattcgata
     6661 gcttggctgc aggtcgacct cgaggatgtg cgcggaaccc ctatttgttt atttttctaa
     6721 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     6781 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     6841 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     6901 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     6961 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     7021 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     7081 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     7141 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     7201 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     7261 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     7321 cgtgacacca cgatgcctgc agcaatggca acaacgttgc gcaaactatt aactggcgaa
     7381 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     7441 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     7501 ggtgagcgtg gctctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     7561 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     7621 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     7681 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     7741 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     7801 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     7861 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     7921 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     7981 gtgtagccgt agttaggcca ccacttcaag aa
//
