LOCUS       sequence-115449-        6050 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses malE-35-H-6 in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6050)
  AUTHORS   Tang NC, Chilkoti A
  TITLE     Combinatorial codon scrambling enables scalable gene synthesis and 
            amplification of repetitive proteins.
  JOURNAL   Nat Mater. 2016 Jan 4. doi: 10.1038/nmat4521.
  PUBMED    26726995
REFERENCE   2  (bases 1 to 6050)
  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: "Nat Mater. 
            2016 Jan 4. doi: 10.1038/nmat4521."
COMMENT     Sequence Label: pMAL-c5X-35-H-6
FEATURES             Location/Qualifiers
     source          1..6050
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..80
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             81..1163
                     /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     complement(100..119)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1406..1411,1412..1427,1428..1434)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1442..1458
                     /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)."
     primer_bind     1447..1469
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1528..2628
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2602..2625
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2677..2688
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3103..3120
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3136..3222
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3314..3341
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3369..3459
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3460..3528,3529..4320)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3678..3697)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4408..4996
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4897..4916
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5150..5167
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     5350..5372
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5366..5557)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
       61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatttcggta gtgggatacg
      901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
      961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1501 gcaattgacc aacaaggacc atagattatg aaaatcgaag aaggtaaact ggtaatctgg
     1561 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     1621 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     1681 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     1741 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     1801 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     1861 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     1921 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     1981 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     2041 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     2101 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     2161 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     2221 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     2281 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     2341 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     2401 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     2461 ttggtgaaag atccgcgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg
     2521 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     2581 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg
     2641 aacaacaaca acaataacaa taacaacaac ctcgggatcg agggaaggtc ttccggtgca
     2701 gcagcgcaag cagctcaggc tcaggctgca gcggaagcag ccgcccaagc tgcccaggca
     2761 caggcggcag cacaggccgc acaggctcaa gctgctgcgg aggcagctgc gcaagcggca
     2821 caagctcagg cagcggctca agccgctcag gcccaagcag ctgcagaggc tgctgcacag
     2881 gcagctcaag cacaggctgc cgcacaagct gcacaagcac aagcagcagc agaagctgcg
     2941 gcgcaagctg ctcaggcaca agcggctgcc caagcggccc aggcgcaggc agcagctgaa
     3001 gctgcagcac aagccgcgca agcccaagcc gcagcccagg ctgctcaagc tcaagcggcg
     3061 gccgaagctg ccgctcaagc agcccaagct caaggctcgg gacatcatca ccaccatcat
     3121 taataatgaa agcttcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt
     3181 atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg
     3241 aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg
     3301 catcaaatta agcagaaggc catcctgacg gatggccttt ttgcgtttct acaaactctt
     3361 tcggtccgtt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     3421 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     3481 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     3541 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     3601 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt cccaatgatg
     3661 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     3721 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     3781 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     3841 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     3901 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     3961 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     4021 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     4081 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     4141 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     4201 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     4261 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     4321 ctgtcagacc aagtttactc atatatactt tagattgatt tccttaggac tgagcgtcaa
     4381 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     4441 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     4501 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     4561 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     4621 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     4681 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     4741 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     4801 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     4861 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     4921 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     4981 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     5041 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     5101 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     5161 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     5221 ttcacaccgc atataaggtg cactgtgact gggtcatggc tgcgccccga cacccgccaa
     5281 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     5341 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     5401 ggcagctgcg gtaaagctca tcagcgtggt cgtgcagcga ttcacagatg tctgcctgtt
     5461 catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc
     5521 gggccatgtt aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat
     5581 ttctgttcat gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta
     5641 ctgatgatga acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga
     5701 tgcggcggga ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg
     5761 taggtgttcc acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc
     5821 agggcgctga cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg
     5881 ttgttgctca ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg
     5941 gtgattcatt ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca
     6001 ggagcacgat catgcgcacc cgtggccagg acccaacgct gcccgaaatt
//
