LOCUS       sequence-385724-        6012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression/purification of 6xHis-TEV-MlaD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6012)
  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 6012)
  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: pBEL1160
FEATURES             Location/Qualifiers
     source          1..6012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      30..485
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(117..136)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     327..348
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(560..593)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        672..776
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(777..845,846..1637)
                     /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(995..1014)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1808..2396
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2297..2316
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2550..2567
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2582..2721
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2647..2666)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2807..2829
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2823..3014)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3823..4905)
                     /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     4867..4886
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4906..4983)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5189..5208
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5292..5311
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5292..5310
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5311..5335
                     /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             5384..5401
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5944..5962)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5958..6005
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 gaggaactat atccggattg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc
       61 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
      121 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
      181 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
      241 tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
      301 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
      361 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
      421 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac
      481 aatttaggtg gcacttttcg gggaacgacg acctgcagcc aagctagctt ggctggacgt
      541 aaactcctct tcagacctaa taacttcgta tagcatacat tatacgaagt tatattaagg
      601 gttattgaat atgatcggaa ttgggctgca ggaattcgat agcttggctg caggtcgacc
      661 tcgaggatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
      721 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
      781 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
      841 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
      901 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
      961 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     1021 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     1081 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     1141 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     1201 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     1261 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     1321 cagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     1381 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     1441 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt ggctctcgcg
     1501 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     1561 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     1621 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     1681 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     1741 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     1801 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     1861 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     1921 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     1981 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2041 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2101 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2161 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2221 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2281 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2341 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2401 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2461 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2521 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2581 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     2641 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     2701 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     2761 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     2821 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     2881 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     2941 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     3001 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     3061 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     3121 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     3181 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     3241 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     3301 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     3361 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     3421 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     3481 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     3541 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     3601 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     3661 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     3721 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     3781 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     3841 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     3901 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     3961 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     4021 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     4081 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     4141 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4201 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4261 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4321 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4381 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4441 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     4501 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4561 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4621 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     4681 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     4741 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     4801 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     4861 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     4921 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     4981 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     5041 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     5101 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     5161 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     5221 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     5281 tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     5341 agaaataatt ttgtttaact ttggcccagg agatatacat atgcatcacc accatcacca
     5401 tgaaaatctg tattttcaga cgtccatacg tactgaaccg acctacacgc tttatgcgac
     5461 gttcgataac attggcggcc tgaaagcgcg ctctccggtc agtattggtg gcgttgttgt
     5521 gggtcgggtg gcggatatta cgctggaccc gaaaacctat ctgccgcgcg taacgctgga
     5581 aattgaacaa cgttataacc acattccaga taccagttcg ctgagcattc gtacttccgg
     5641 cctgctgggg gaacaatatc tggcattaaa cgtcggtttt gaagacccgg aactggggac
     5701 tgctatcctg aaggatggcg atacaattca ggacactaag tctgcgatgg tgctggaaga
     5761 tctcattggt cagttccttt acggtagtaa aggcgatgac aataagaata gtggcgatgc
     5821 gccagctgct gcgccaggta ataatgaaac cactgaacct gtgggtacaa cgaaataaga
     5881 gagaccggcc ggccgatccg gctgctaaca aagcccgaaa ggaagctgag ttggctgctg
     5941 ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt
     6001 ttttgctgaa ag
//
