LOCUS       sequence-69778-e        6552 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 6552)
  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 6552)
  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 TEV cloning vector with BioB...
FEATURES             Location/Qualifiers
     source          1..6552
                     /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     1474..1491
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             1611..1628
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1638..2738
                     /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     2712..2735
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2793..2813
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     polyA_signal    3039..3173
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3076..3095)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3130..3149
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  3218..3383
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      3567..4022
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3654..3673)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3864..3885
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4049..4153
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4154..4222,4223..5014)
                     /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(4372..4391)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5185..5773
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5674..5693
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5927..5944
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(6028..6047)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6188..6210
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6204..6395)
                     /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 gccgcttaat taaaaatgat aaccatctcg caaataaata
     1501 agtattttac tgttttcgta acagttttgt aataaaaaaa cctataacta ttccggatta
     1561 ttcataccgt cccaccatcg ggcgcggatc ccggaccgat gggttcttct caccatcacc
     1621 atcaccatgg ttcttctatg aaaatcgaag aaggtaaact ggtaatctgg attaacggcg
     1681 ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat accggaatta
     1741 aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt gcggcaactg
     1801 gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac gctcaatctg
     1861 gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat ccgtttacct
     1921 gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat
     1981 cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa gagatcccgg
     2041 cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac ctgcaagaac
     2101 cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag tatgaaaacg
     2161 gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg ggtctgacct
     2221 tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac tccatcgcag
     2281 aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca
     2341 acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc aagggtcaac
     2401 catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt ccgaacaaag
     2461 agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg gaagcggtta
     2521 ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag ttggcgaaag
     2581 atccacgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc
     2641 cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc
     2701 gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg aacaacaaca
     2761 acaataacaa taacaacaac ctcgggatcg aggaaaacct gtacttccaa tccaatattg
     2821 gaagtggata acggatccga attcgagcgc cgtcgacaag cttgcggcct catgatctcc
     2881 taggggccgc actcgaggca tgcggtacca agcttgtcga gaagtactag aggatcataa
     2941 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     3001 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     3061 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3121 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg atctgatcac
     3181 tgcttgagcc tagcgatcgc cctgcagggg agatccgaac cagataagtg aaatctagtt
     3241 ccaaactatt ttgtcatttt taattttcgt attagcttac gacgctacac ccagttccca
     3301 tctattttgt cactcttccc taaataatcc ttaaaaactc catttccacc cctcccagtt
     3361 cccaactatt ttgtccgccc acagcggggc atttttcttc ctgttatgtt tttaatcaaa
     3421 catcctgcca actccatgtg acaaaccgtc atcttcggct actttttctc tgtcacagaa
     3481 tgaaaatttt tctgtcatct cttcgttatt aatgtttgta attgactgaa tatcaacgct
     3541 tatttgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg
     3601 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     3661 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     3721 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     3781 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     3841 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     3901 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     3961 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaactattaa cgtttacaat
     4021 ttcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     4081 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aatactattg
     4141 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     4201 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     4261 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     4321 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     4381 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     4441 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     4501 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     4561 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     4621 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     4681 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     4741 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     4801 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     4861 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     4921 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     4981 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     5041 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     5101 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     5161 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     5221 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     5281 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt
     5341 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     5401 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     5461 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     5521 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     5581 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     5641 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     5701 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     5761 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     5821 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     5881 ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag
     5941 cgaggaagcg gaagaggcgc gccatgcggt attttctcct tacgcatctg tgcggtattt
     6001 cacaccgcat atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     6061 gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca
     6121 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     6181 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg
     6241 cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca
     6301 tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg
     6361 gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt
     6421 ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact
     6481 gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg
     6541 cggcgggacc ag
//
