LOCUS       sequence-85605-e        6606 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6606)
  AUTHORS   Gradia SD, Ishida JP, Tsai MS, Jeans C, Tainer JA, Fuss JO
  TITLE     MacroBac: New Technologies for Robust and Efficient Large-Scale 
            Production of Recombinant Multiprotein Complexes.
  JOURNAL   Methods Enzymol. 2017;592:1-26. doi: 10.1016/bs.mie.2017.03.008. 
            Epub 2017 May 15.
  PUBMED    28668116
REFERENCE   2  (bases 1 to 6606)
  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: "Methods 
            Enzymol."; date: "2017"; volume: "592:1-26. doi"; pages: " 
            10.1016/bs.mie.2017.03.008. Epub 2017 May 15"
COMMENT     Sequence Label: pFastBac His6 MBP Asn10 TEV cloning vector wit...
FEATURES             Location/Qualifiers
     source          1..6606
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     mobile_element  68..292
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(359..892)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLWSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(1062..1067,1068..1084,1085..1090))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     primer_bind     1499..1516
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             1636..1653
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1663..2763
                     /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
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2737..2760
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2818..2838
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     polyA_signal    3064..3198
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3101..3120)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3155..3174
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  3272..3437
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      3621..4076
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3708..3727)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3918..3939
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4103..4207
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4208..4276,4277..5068)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4426..4445)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5239..5827
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5728..5747
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5981..5998
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(6082..6101)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6242..6264
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6258..6449)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gaccagccgc gtaacctggc aaaatcggtt acggttgagt aataaatgga tgccctgcgt
       61 aagcgggtgt gggcggacaa taaagtctta aactgaacaa aatagatcta aactatgaca
      121 ataaagtctt aaactagaca gaatagttgt aaactgaaat cagtccagtt atgctgtgaa
      181 aaagcatact ggacttttgt tatggctaaa gcaaactctt cattttctga agtgcaaatt
      241 gcccgtcgta ttaaagaggg gcgtggccaa gggcatggta aagactatat tcgcggcgtt
      301 gtgacaattt accgaacaac tccgcggccg ggaagccgat ctcggcttga acgaattgtt
      361 aggtggcggt acttgggtcg atatcaaagt gcatcacttc ttcccgtatg cccaactttg
      421 tatagagagc cactgcggga tcgtcaccgt aatctgcttg cacgtagatc acataagcac
      481 caagcgcgtt ggcctcatgc ttgaggagat tgatgagcgc ggtggcaatg ccctgcctcc
      541 ggtgctcgcc ggagactgcg agatcataga tatagatctc actacgcggc tgctcaaact
      601 tgggcagaac gtaagccgcg agagcgccaa caaccgcttc ttggtcgaag gcagcaagcg
      661 cgatgaatgt cttactacgg agcaagttcc cgaggtaatc ggagtccggc tgatgttggg
      721 agtaggtggc tacgtctccg aactcacgac cgaaaagatc aagagcagcc cgcatggatt
      781 tgacttggtc agggccgagc ctacatgtgc gaatgatgcc catacttgag ccacctaact
      841 ttgttttagg gcgactgccc tgctgcgtaa catcgttgct gctccataac atcaaacatc
      901 gacccacggc gtaacgcgct tgctgcttgg atgcccgagg catagactgt acaaaaaaac
      961 agtcataaca agccatgaaa accgccactg cgccgttacc accgctgcgt tcggtcaagg
     1021 ttctggacca gttgcgtgag cgcatacgct acttgcatta cagtttacga accgaacagg
     1081 cttatgtcaa ctgggttcgt gccttcatcc gtttccacgg tgtgcgtcac ccggcaacct
     1141 tgggcagcag cgaagtcgag gcatttctgt cctggctggc gaacgagcgc aaggtttcgg
     1201 tctccacgca tcgtcaggca ttggcggcct tgctgttctt ctacggcaag gtgctgtgca
     1261 cggatctgcc ctggcttcag gagatcggaa gacctcggcc gtcgcggcgc ttgccggtgg
     1321 tgctgacccc ggatgaagtg gttcgcatcc tcggttttct ggaaggcgag catcgtttgt
     1381 tcgcccagga ctctagctat agttctagtg gttggctacg tatactccgg actattaata
     1441 gatcatggag ataattagcg gccgcgttta aacatccacc atttgatgga gataattaaa
     1501 atgataacca tctcgcaaat aaataagtat tttactgttt tcgtaacagt tttgtaataa
     1561 aaaaacctat aactattccg gattattcat accgtcccac catcgggcgc ggatcccgga
     1621 ccgatgggtt cttctcacca tcaccatcac catggttctt ctatgaaaat cgaagaaggt
     1681 aaactggtaa tctggattaa cggcgataaa ggctataacg gtctcgctga agtcggtaag
     1741 aaattcgaga aagataccgg aattaaagtc accgttgagc atccggataa actggaagag
     1801 aaattcccac aggttgcggc aactggcgat ggccctgaca ttatcttctg ggcacacgac
     1861 cgctttggtg gctacgctca atctggcctg ttggctgaaa tcaccccgga caaagcgttc
     1921 caggacaagc tgtatccgtt tacctgggat gccgtacgtt acaacggcaa gctgattgct
     1981 tacccgatcg ctgttgaagc gttatcgctg atttataaca aagatctgct gccgaacccg
     2041 ccaaaaacct gggaagagat cccggcgctg gataaagaac tgaaagcgaa aggtaagagc
     2101 gcgctgatgt tcaacctgca agaaccgtac ttcacctggc cgctgattgc tgctgacggg
     2161 ggttatgcgt tcaagtatga aaacggcaag tacgacatta aagacgtggg cgtggataac
     2221 gctggcgcga aagcgggtct gaccttcctg gttgacctga ttaaaaacaa acacatgaat
     2281 gcagacaccg attactccat cgcagaagct gcctttaata aaggcgaaac agcgatgacc
     2341 atcaacggcc cgtgggcatg gtccaacatc gacaccagca aagtgaatta tggtgtaacg
     2401 gtactgccga ccttcaaggg tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt
     2461 attaacgccg ccagtccgaa caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg
     2521 actgatgaag gtctggaagc ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag
     2581 tcttacgagg aagagttggc gaaagatcca cgtattgccg ccactatgga aaacgcccag
     2641 aaaggtgaaa tcatgccgaa catcccgcag atgtccgctt tctggtatgc cgtgcgtact
     2701 gcggtgatca acgccgccag cggtcgtcag actgtcgatg aagccctgaa agacgcgcag
     2761 actaattcga gctcgaacaa caacaacaat aacaataaca acaacctcgg gatcgaggaa
     2821 aacctgtact tccaatccaa tattggaagt ggataacgga tccgaattcg agcgccgtcg
     2881 acaagcttgc ggcctcatga tctcctaggg gccgcactcg aggcatgcgg taccaagctt
     2941 gtcgagaagt actagaggat cataatcagc cataccacat ttgtagaggt tttacttgct
     3001 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
     3061 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     3121 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3181 tcttatcatg tctggatctg atcactgctt gagcctagaa acatccacca tttaaatgga
     3241 gttaggttta aacgcctgca ggggagatcc gaaccagata agtgaaatct agttccaaac
     3301 tattttgtca tttttaattt tcgtattagc ttacgacgct acacccagtt cccatctatt
     3361 ttgtcactct tccctaaata atccttaaaa actccatttc cacccctccc agttcccaac
     3421 tattttgtcc gcccacagcg gggcattttt cttcctgtta tgtttttaat caaacatcct
     3481 gccaactcca tgtgacaaac cgtcatcttc ggctactttt tctctgtcac agaatgaaaa
     3541 tttttctgtc atctcttcgt tattaatgtt tgtaattgac tgaatatcaa cgcttatttg
     3601 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     3661 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     3721 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     3781 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     3841 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3901 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     3961 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     4021 gctgatttaa caaaaattta acgcgaattt taacaaacta ttaacgttta caatttcagg
     4081 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4141 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaatact attgaaaaag
     4201 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     4261 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     4321 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     4381 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     4441 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     4501 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     4561 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     4621 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     4681 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     4741 cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     4801 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     4861 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     4921 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     4981 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     5041 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     5101 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     5161 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     5221 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5281 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5341 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     5401 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5461 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5521 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5581 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5641 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5701 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5761 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5821 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5881 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     5941 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     6001 agcggaagag gcgcgccatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     6061 gcatatatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatac
     6121 actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc aacacccgct
     6181 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     6241 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg
     6301 cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg
     6361 tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg
     6421 ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg atttctgttc
     6481 atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt tactgatgat
     6541 gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg gatgcggcgg
     6601 gaccag
//
