LOCUS       sequence-224676-        7144 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal HIS6-MBP tag, cleavable with
            3C, and C-terminal SNAP tag, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7144)
  AUTHORS   Lemaitre RP, Bogdanova A, Borgonovo B, Woodruff JB, Drechsel DN
  TITLE     FlexiBAC: a versatile, open-source baculovirus vector system for 
            protein expression, secretion, and proteolytic processing.
  JOURNAL   BMC Biotechnol. 2019 Mar 29;19(1):20. doi: 
            10.1186/s12896-019-0512-z.
  PUBMED    30925874
REFERENCE   2  (bases 1 to 7144)
  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: "BMC 
            Biotechnol."; date: "2019-03-29"; volume: "19(1):20. doi"; pages: " 
            10.1186/s12896-019-0512-z"
COMMENT     Sequence Label: pOCC177
FEATURES             Location/Qualifiers
     source          1..7144
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        79..170
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     95..112
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     regulatory      212..221
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             230..247
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             260..1360
                     /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     1334..1357
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1415..1438
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1547..1852
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1731..1750
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1908..1928
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1929..2474
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     polyA_signal    2610..2744
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2647..2666)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2701..2720
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2817..3189
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3541..3558)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3651..4239)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3731..3750)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4410..5201,5202..5270))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5033..5052
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5272..5376)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5677..5699
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5691..5708
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5692..5708
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5719..5738
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5719..5737
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6348..6472
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6510..7115)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
ORIGIN
        1 agctgccacc attttgttcg cccaggactc tagctatagt tctagtggtt ggctacgtat
       61 actccggaat attaatagat catggagata attaaaatga taaccatctc gcaaataaat
      121 aagtatttta ctgttttcgt aacagttttg taataaaaaa acctataaat attccggatt
      181 attcataccg tcccaccatc gggcgcggat cgccaccatg ggcagcagcc atcaccatca
      241 ccatcacagc agcggcagaa tgaaaatcga agaaggtaaa ctggtaatct ggattaacgg
      301 cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat
      361 taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac
      421 tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc
      481 tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac
      541 ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt
      601 atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc
      661 ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga
      721 accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa
      781 cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac
      841 cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc
      901 agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc
      961 caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca
     1021 accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa
     1081 agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt
     1141 taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggtgaa
     1201 agatccgcgt attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat
     1261 cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg
     1321 tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa
     1381 caacaataac aataacaaca acagcagcgg cagactggaa gttctgttcc aggggccggc
     1441 ggccgcaagc gctatggcca caacctgtta cattgcacaa gataaaaata tatcatcacg
     1501 aacagtaaaa ctgtctgctt acataaacag taatacaagg ggtgttatgc agtttaaggt
     1561 ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga gtgatattat
     1621 tgacacgccc gggcgacgga tggtgatccc cctggccagt gcccgtctgc tgtcagataa
     1681 agtctcccgt gaactttacc cggtggtgca tatcggggat gaaagctggc gcatgatgac
     1741 caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg atctcagcca
     1801 ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat aaatgtcagg
     1861 ctcccttata cacagccagt ggcgcgccgg gatccagcag cggcagagaa aacttgtatt
     1921 tccagggcat ggacaaagac tgcgaaatga agcgcaccac cctggatagc cctctgggca
     1981 agctggaact gtctgggtgc gaacagggcc tgcaccgtat catcttcctg ggcaaaggaa
     2041 catctgccgc cgacgccgtg gaagtgcctg ccccagccgc cgtgctgggc ggaccagagc
     2101 cactgatgca ggccaccgcc tggctcaacg cctactttca ccagcctgag gccatcgagg
     2161 agttccctgt gccagccctg caccacccag tgttccagca ggagagcttt acccgccagg
     2221 tgctgtggaa actgctgaaa gtggtgaagt tcggagaggt catcagctac agccacctgg
     2281 ccgccctggc cggcaatccc gccgccaccg ccgccgtgaa aaccgccctg agcggaaatc
     2341 ccgtgcccat tctgatcccc tgccaccggg tggtgcaggg cgacctggac gtggggggct
     2401 acgagggcgg gctcgccgtg aaagagtggc tgctggccca cgagggccac agactgggca
     2461 agcctgggct gggttaatag aagcttgtcg agaagtacta gaggatcata atcagccata
     2521 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     2581 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     2641 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2701 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatctgatca ctgcttgagc
     2761 ctaggagatc cgaaccagat aagtgaaagc ctgcatccgc agatcttaat taataaaaca
     2821 cgatacattg ttattagtac atttattaag cgctagattc tgtgcgttgt tgatttacag
     2881 acaattgttg tacgtatttt aataattcat taaatttata atctttaggg tggtatgtta
     2941 gagcgaaaat caaatgattt tcagcgtctt tatatctgaa tttaaatatt aaatcctcaa
     3001 tagatttgta aaataggttt cgattagttt caaacaaggg ttgtttttcc gaaccgatgg
     3061 ctggactatc taatggattt tcgctcaacg ccacaaaact tgccaaatct tgtagcagca
     3121 atctagcttt gtcgatattc gtttgtgttt tgttttgtaa taaaggttcg acgtcgttca
     3181 aaatattatg cgcttttgta tttctttcat cactgtcgtt agtgtacaat tgactcgacg
     3241 taaacacgtt aaataacgct tggacatatt taacatcggg cgtgttagct ttattaggcc
     3301 gattatcgtc gtcgtcccaa ccctcgtcgt tagaagttgc ttccgaagac gattttgcca
     3361 tagccacacg acgcctatta attgtgtcgg ctaacacgtc cgcgatcaaa tttgtagttg
     3421 agctttttgg gcctcatggg ccttccgctc actgcccgct ttccagtcgg gaaacctgtc
     3481 gtgccagctg cattaacatg gtcatagctg tttccttgcg tattgggcgc tctccgcttc
     3541 ctcgctcact gactcgctgc gctcggtcgt tcgggtaaag cctggggtgc ctaatgagca
     3601 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3661 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3721 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3781 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3841 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3901 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3961 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     4021 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     4081 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     4141 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4201 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4261 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4321 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4381 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4441 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4501 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4561 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4621 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4681 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg
     4741 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4801 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4861 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4921 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4981 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc
     5041 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     5101 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     5161 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5221 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat tactcttcct
     5281 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     5341 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     5401 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     5461 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     5521 atagggttga gtggccgcta cagggcgctc ccattcgcca ttcaggctgc gcaactgttg
     5581 ggaagggcgt ttcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     5641 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     5701 cggccagtga gcgcgacgta atacgactca ctatagggcg aattggcgga aggccgtcaa
     5761 ggccgtgacg ttaaaactat taagccatcc aatcgaccgt tagtcgaatc aggaccgctg
     5821 gtgcgagaag ccgcgaagta tggcgaatgc atcgtataac gtgtggagtc cgctcattag
     5881 agcgtcatgt ttagacaaga aagctacata tttaattgat cccgatgatt ttattgataa
     5941 attgacccta actccataca cggtattcta caatggcggg gttttggtca aaatttccgg
     6001 actgcgattg tacatgctgt taacggctcc gcccactatt aatgaaatta aaaattccaa
     6061 ttttaaaaaa cgcagcaaga gaaacatttg tatgaaagaa tgcgtagaag gaaagaaaaa
     6121 tgtcgtggac atgctgaaca acaagattaa tatgcctccg tgtataaaaa aaatattgaa
     6181 cgatttgaaa gaaaacaatg taccgcgcgg cggtatgtac aggaagaggt ttatactaaa
     6241 ctgttacatt gcaaacgtgg tttcgtgtgc caagtgtgaa aaccgatgtt taatcaaggc
     6301 tctgacgcat ttctacaacc acgactccaa gtgtgtgggt gaagtcatgc atcttttaat
     6361 caaatcccaa gatgtgtata aaccaccaaa ctgccaaaaa atgaaaactg tggacaagct
     6421 ctgtccgttt gctggcaact gcaagggtct caatcctatt tgtaattatt gaataataaa
     6481 acaattataa atgctaaatt tgttttttat taacgataca aaccaaacgc aacaagaaca
     6541 tttgtagtat tatctataat tgaaaacgcg tagttataat cgctgaggta atatttaaaa
     6601 tcattttcaa atgattcaca gttaatttgc gacaatataa ttttattttc acataaacta
     6661 gacgccttgt cgtcttcttc ttcgtattcc ttctcttttt catttttctc ctcataaaaa
     6721 ttaacatagt tattatcgta tccatatatg tatctatcgt atagagtaaa ttttttgttg
     6781 tcataaatat atatgtcttt tttaatgggg tgtatagtac cgctgcgcat agtttttctg
     6841 taatttacaa cagtgctatt ttctggtagt tcttcggagt gtgttgcttt aattattaaa
     6901 tttatataat caatgaattt gggatcgtcg gttttgtaca atatgttgcc ggcatagtac
     6961 gcagcttctt ctagttcaat tacaccattt tttagcagca ccggattaac ataactttcc
     7021 aaaatgttgt acgaaccgtt aaacaaaaac agttcacctc ccttttctat actattgtct
     7081 gcgagcagtt gtttgttgtt aaaaataaca gccattgtaa tgagacgcac aaactaatat
     7141 caca
//
