LOCUS       sequence-385719-        5883 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression/purification of 6xHis-TEV-MlaDdelta141-183.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5883)
  AUTHORS   MacRae MR, Puvanendran D, Haase MAB, Coudray N, Kolich L, Lam C, 
            Baek M, Bhabha G, Ekiert DC
  TITLE     Protein-protein interactions in the Mla lipid transport system 
            probed by computational structure prediction and deep mutational 
            scanning.
  JOURNAL   J Biol Chem. 2023 Apr 24:104744. doi: 10.1016/j.jbc.2023.104744.
  PUBMED    37100290
REFERENCE   2  (bases 1 to 5883)
  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 Biol 
            Chem. 2023 Apr 24:104744. doi: 10.1016/j.jbc.2023.104744."
COMMENT     Sequence Label: pBEL1161
FEATURES             Location/Qualifiers
     source          1..5883
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      28..483
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(115..134)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     325..346
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(558..591)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        670..774
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(775..843,844..1635)
                     /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(993..1012)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1806..2394
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2295..2314
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2548..2565
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2580..2719
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2645..2664)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2805..2827
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2821..3012)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3821..4903)
                     /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     4865..4884
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4904..4981)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5187..5206
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5290..5309
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5290..5308
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5309..5333
                     /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             5382..5399
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5813..5831)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5827..5874
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
       61 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
      121 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
      181 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
      241 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
      301 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
      361 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
      421 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
      481 tttaggtggc acttttcggg gaacgacgac ctgcagccaa gctagcttgg ctggacgtaa
      541 actcctcttc agacctaata acttcgtata gcatacatta tacgaagtta tattaagggt
      601 tattgaatat gatcggaatt gggctgcagg aattcgatag cttggctgca ggtcgacctc
      661 gaggatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
      721 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
      781 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
      841 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      901 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      961 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     1021 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1081 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1141 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1201 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     1261 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca
     1321 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     1381 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     1441 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg ctctcgcggt
     1501 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     1561 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     1621 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     1681 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     1741 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     1801 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     1861 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     1921 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     1981 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     2041 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2101 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2161 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     2221 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     2281 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     2341 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     2401 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     2461 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     2521 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     2581 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc
     2641 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     2701 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     2761 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     2821 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     2881 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     2941 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     3001 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     3061 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     3121 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     3181 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     3241 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     3301 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     3361 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     3421 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     3481 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
     3541 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     3601 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
     3661 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     3721 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     3781 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc tcactgcccg ctttccagtc
     3841 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     3901 gcgtattggg cgccagggtg gtttttcttt tcaccagtga gacgggcaac agctgattgc
     3961 ccttcaccgc ctggccctga gagagttgca gcaagcggtc cacgctggtt tgccccagca
     4021 ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata acatgagctg tcttcggtat
     4081 cgtcgtatcc cactaccgag atatccgcac caacgcgcag cccggactcg gtaatggcgc
     4141 gcattgcgcc cagcgccatc tgatcgttgg caaccagcat cgcagtggga acgatgccct
     4201 cattcagcat ttgcatggtt tgttgaaaac cggacatggc actccagtcg ccttcccgtt
     4261 ccgctatcgg ctgaatttga ttgcgagtga gatatttatg ccagccagcc agacgcagac
     4321 gcgccgagac agaacttaat gggcccgcta acagcgcgat ttgctggtga cccaatgcga
     4381 ccagatgctc cacgcccagt cgcgtaccgt cttcatggga gaaaataata ctgttgatgg
     4441 gtgtctggtc agagacatca agaaataacg ccggaacatt agtgcaggca gcttccacag
     4501 caatggcatc ctggtcatcc agcggatagt taatgatcag cccactgacg cgttgcgcga
     4561 gaagattgtg caccgccgct ttacaggctt cgacgccgct tcgttctacc atcgacacca
     4621 ccacgctggc acccagttga tcggcgcgag atttaatcgc cgcgacaatt tgcgacggcg
     4681 cgtgcagggc cagactggag gtggcaacgc caatcagcaa cgactgtttg cccgccagtt
     4741 gttgtgccac gcggttggga atgtaattca gctccgccat cgccgcttcc actttttccc
     4801 gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg ggaaacggtc tgataagaga
     4861 caccggcata ctctgcgaca tcgtataacg ttactggttt cacattcacc accctgaatt
     4921 gactctcttc cgggcgctat catgccatac cgcgaaaggt tttgcgccat tcgatggtgt
     4981 ccgggatctc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     5041 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     5101 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     5161 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     5221 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     5281 ccgcgaaatt aatacgactc actatagggg aattgtgagc ggataacaat tcccctctag
     5341 aaataatttt gtttaacttt ggcccaggag atatacatat gcatcaccac catcaccatg
     5401 aaaatctgta ttttcagacg tccatacgta ctgaaccgac ctacacgctt tatgcgacgt
     5461 tcgataacat tggcggcctg aaagcgcgct ctccggtcag tattggtggc gttgttgtgg
     5521 gtcgggtggc ggatattacg ctggacccga aaacctatct gccgcgcgta acgctggaaa
     5581 ttgaacaacg ttataaccac attccagata ccagttcgct gagcattcgt acttccggcc
     5641 tgctggggga acaatatctg gcattaaacg tcggttttga agacccggaa ctggggactg
     5701 ctatcctgaa ggatggcgat acaattcagg acactaagtc tgcgtaagag agaccggccg
     5761 gccgatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     5821 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
     5881 gga
//
