LOCUS       sequence-385684-        6126 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression/purification of 6xHis-TEV-MlaC .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6126)
  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 6126)
  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: pBEL1203
FEATURES             Location/Qualifiers
     source          1..6126
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      33..488
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(120..139)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     330..351
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(563..596)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        675..779
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(780..848,849..1640)
                     /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(998..1017)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1811..2399
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2300..2319
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2553..2570
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2585..2724
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2650..2669)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2810..2832
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2826..3017)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3826..4908)
                     /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     4870..4889
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4909..4986)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5192..5211
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5295..5314
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5295..5313
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5314..5338
                     /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             5387..5404
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             5405..5425
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQA"
     primer_bind     complement(6061..6079)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6075..6122
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 aaggaggaac tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
       61 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
      121 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
      181 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
      241 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
      301 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
      361 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
      421 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
      481 tacaatttag gtggcacttt tcggggaacg acgacctgca gccaagctag cttggctgga
      541 cgtaaactcc tcttcagacc taataacttc gtatagcata cattatacga agttatatta
      601 agggttattg aatatgatcg gaattgggct gcaggaattc gatagcttgg ctgcaggtcg
      661 acctcgagga tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
      721 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
      781 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
      841 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
      901 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
      961 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     1021 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1081 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1141 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1201 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1261 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     1321 ctgcagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     1381 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     1441 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggctctc
     1501 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     1561 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     1621 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     1681 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     1741 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     1801 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     1861 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     1921 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     1981 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2041 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2101 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2161 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2221 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2281 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     2341 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     2401 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     2461 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     2521 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     2581 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaatggtg
     2641 cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac tccgctatcg
     2701 ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga cgcgccctga
     2761 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     2821 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca
     2881 tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc cagctcgttg
     2941 agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt aagggcggtt
     3001 ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat gggggtaatg
     3061 ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga acatgcccgg
     3121 ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga ccagagaaaa
     3181 atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc acagggtagc
     3241 cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga cttccgcgtt
     3301 tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca ggtcgcagac
     3361 gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca
     3421 gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat catgcgcacc
     3481 cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga
     3541 ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg
     3601 atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc
     3661 acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg
     3721 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     3781 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     3841 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     3901 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     3961 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     4021 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     4081 ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg actcggtaat
     4141 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     4201 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     4261 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     4321 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     4381 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     4441 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     4501 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     4561 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     4621 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     4681 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     4741 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     4801 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     4861 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     4921 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     4981 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     5041 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     5101 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     5161 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     5221 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatct
     5281 cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata acaattcccc
     5341 tctagaaata attttgttta actttggccc aggagatata catatgcatc accaccatca
     5401 ccatgaaaat ctgtattttc aggcagacca gaccaatccg tataagctga tggacgaggc
     5461 ggcgcagaaa acgttcgatc gcctgaagaa tgagcaaccg caaattcggg ccaacccgga
     5521 ttatctgcgt accattgttg atcaggaact gctgccatac gtacaggtga aatacgccgg
     5581 tgcgctggtg ctgggccagt attacaagag tgcgacccct gctcaacgtg aagcctactt
     5641 tgccgctttc cgtgagtacc tgaagcaggc ttacggtcag gcgctggcga tgtatcacgg
     5701 tcaaacctat cagattgcgc cagaacagcc gctgggcgat aaaaccattg tgcctattcg
     5761 cgttaccatt attgacccga atggccgtcc gccggtgcgt ctggacttcc agtggcgtaa
     5821 aaactcccag acgggcaatt ggcaggctta cgacatgatt gctgaaggcg tcagtatgat
     5881 caccaccaaa caaaacgagt ggggaacgct gctgcgtacc aaaggtatcg acggcctgac
     5941 tgcgcaactg aaatcgattt ctcaacagaa aatcactctg gaagagaaaa aataagagag
     6001 accggccggc cgatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     6061 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     6121 tgctga
//
