LOCUS       sequence-224674-        7309 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 monomeric GFP, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7309)
  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 7309)
  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: pOCC119
FEATURES             Location/Qualifiers
     source          1..7309
                     /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             224..241
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             254..1354
                     /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     1328..1351
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1409..1432
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1541..1846
                     /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     1725..1744
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1902..1922
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(1923..1925,1926..1928,1929..2642)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1968..1989)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2229..2248)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2576..2597
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2775..2909
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2812..2831)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2866..2885
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2982..3354
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3706..3723)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3816..4404)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3896..3915)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4575..5366,5367..5435))
                     /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     5198..5217
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5437..5541)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5842..5864
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5856..5873
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5857..5873
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5884..5903
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5884..5902
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6513..6637
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6675..7280)
                     /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 ggccatcacc atcaccatca
      241 cagcagcggc agaatgaaaa tcgaagaagg taaactggta atctggatta acggcgataa
      301 aggctataac ggtctcgctg aagtcggtaa gaaattcgag aaagataccg gaattaaagt
      361 caccgttgag catccggata aactggaaga gaaattccca caggttgcgg caactggcga
      421 tggccctgac attatcttct gggcacacga ccgctttggt ggctacgctc aatctggcct
      481 gttggctgaa atcaccccgg acaaagcgtt ccaggacaag ctgtatccgt ttacctggga
      541 tgccgtacgt tacaacggca agctgattgc ttacccgatc gctgttgaag cgttatcgct
      601 gatttataac aaagatctgc tgccgaaccc gccaaaaacc tgggaagaga tcccggcgct
      661 ggataaagaa ctgaaagcga aaggtaagag cgcgctgatg ttcaacctgc aagaaccgta
      721 cttcacctgg ccgctgattg ctgctgacgg gggttatgcg ttcaagtatg aaaacggcaa
      781 gtacgacatt aaagacgtgg gcgtggataa cgctggcgcg aaagcgggtc tgaccttcct
      841 ggttgacctg attaaaaaca aacacatgaa tgcagacacc gattactcca tcgcagaagc
      901 tgcctttaat aaaggcgaaa cagcgatgac catcaacggc ccgtgggcat ggtccaacat
      961 cgacaccagc aaagtgaatt atggtgtaac ggtactgccg accttcaagg gtcaaccatc
     1021 caaaccgttc gttggcgtgc tgagcgcagg tattaacgcc gccagtccga acaaagagct
     1081 ggcaaaagag ttcctcgaaa actatctgct gactgatgaa ggtctggaag cggttaataa
     1141 agacaaaccg ctgggtgccg tagcgctgaa gtcttacgag gaagagttgg tgaaagatcc
     1201 gcgtattgcc gccactatgg aaaacgccca gaaaggtgaa atcatgccga acatcccgca
     1261 gatgtccgct ttctggtatg ccgtgcgtac tgcggtgatc aacgccgcca gcggtcgtca
     1321 gactgtcgat gaagccctga aagacgcgca gactaattcg agctcgaaca acaacaacaa
     1381 taacaataac aacaacagca gcggcagact ggaagttctg ttccaggggc cggcggccgc
     1441 aagcgctatg gccacaacct gttacattgc acaagataaa aatatatcat cacgaacagt
     1501 aaaactgtct gcttacataa acagtaatac aaggggtgtt atgcagttta aggtttacac
     1561 ctataaaaga gagagccgtt atcgtctgtt tgtggatgta cagagtgata ttattgacac
     1621 gcccgggcga cggatggtga tccccctggc cagtgcccgt ctgctgtcag ataaagtctc
     1681 ccgtgaactt tacccggtgg tgcatatcgg ggatgaaagc tggcgcatga tgaccaccga
     1741 tatggccagt gtgccggtct ccgttatcgg ggaagaagtg gctgatctca gccaccgcga
     1801 aaatgacatc aaaaacgcca ttaacctgat gttctgggga atataaatgt caggctccct
     1861 tatacacagc cagtggcgcg ccgggatcca gcagcggcag agaaaacttg tatttccagg
     1921 gcatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1981 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     2041 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     2101 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     2161 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     2221 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     2281 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2341 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     2401 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     2461 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     2521 actacctgag cacccagtcc aaactgagca aagaccccaa cgagaagcgc gatcacatgg
     2581 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     2641 aatagaagct tgtcgagaag tactagagga tcataatcag ccataccaca tttgtagagg
     2701 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     2761 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2821 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     2881 tcatcaatgt atcttatcat gtctggatct gatcactgct tgagcctagg agatccgaac
     2941 cagataagtg aaagcctgca tccgcagatc ttaattaata aaacacgata cattgttatt
     3001 agtacattta ttaagcgcta gattctgtgc gttgttgatt tacagacaat tgttgtacgt
     3061 attttaataa ttcattaaat ttataatctt tagggtggta tgttagagcg aaaatcaaat
     3121 gattttcagc gtctttatat ctgaatttaa atattaaatc ctcaatagat ttgtaaaata
     3181 ggtttcgatt agtttcaaac aagggttgtt tttccgaacc gatggctgga ctatctaatg
     3241 gattttcgct caacgccaca aaacttgcca aatcttgtag cagcaatcta gctttgtcga
     3301 tattcgtttg tgttttgttt tgtaataaag gttcgacgtc gttcaaaata ttatgcgctt
     3361 ttgtatttct ttcatcactg tcgttagtgt acaattgact cgacgtaaac acgttaaata
     3421 acgcttggac atatttaaca tcgggcgtgt tagctttatt aggccgatta tcgtcgtcgt
     3481 cccaaccctc gtcgttagaa gttgcttccg aagacgattt tgccatagcc acacgacgcc
     3541 tattaattgt gtcggctaac acgtccgcga tcaaatttgt agttgagctt tttgggcctc
     3601 atgggccttc cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     3661 acatggtcat agctgtttcc ttgcgtattg ggcgctctcc gcttcctcgc tcactgactc
     3721 gctgcgctcg gtcgttcggg taaagcctgg ggtgcctaat gagcaaaagg ccagcaaaag
     3781 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3841 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3901 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3961 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4021 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4081 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4141 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4201 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4261 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4321 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4381 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4441 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     4501 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4561 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4621 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4681 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     4741 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4801 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4861 aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt
     4921 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4981 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5041 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5101 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5161 cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga
     5221 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5281 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5341 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5401 ggaataaggg cgacacggaa atgttgaata ctcattactc ttcctttttc aatattattg
     5461 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     5521 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     5581 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     5641 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtggc
     5701 cgctacaggg cgctcccatt cgccattcag gctgcgcaac tgttgggaag ggcgtttcgg
     5761 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa
     5821 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgagcgcg
     5881 acgtaatacg actcactata gggcgaattg gcggaaggcc gtcaaggccg tgacgttaaa
     5941 actattaagc catccaatcg accgttagtc gaatcaggac cgctggtgcg agaagccgcg
     6001 aagtatggcg aatgcatcgt ataacgtgtg gagtccgctc attagagcgt catgtttaga
     6061 caagaaagct acatatttaa ttgatcccga tgattttatt gataaattga ccctaactcc
     6121 atacacggta ttctacaatg gcggggtttt ggtcaaaatt tccggactgc gattgtacat
     6181 gctgttaacg gctccgccca ctattaatga aattaaaaat tccaatttta aaaaacgcag
     6241 caagagaaac atttgtatga aagaatgcgt agaaggaaag aaaaatgtcg tggacatgct
     6301 gaacaacaag attaatatgc ctccgtgtat aaaaaaaata ttgaacgatt tgaaagaaaa
     6361 caatgtaccg cgcggcggta tgtacaggaa gaggtttata ctaaactgtt acattgcaaa
     6421 cgtggtttcg tgtgccaagt gtgaaaaccg atgtttaatc aaggctctga cgcatttcta
     6481 caaccacgac tccaagtgtg tgggtgaagt catgcatctt ttaatcaaat cccaagatgt
     6541 gtataaacca ccaaactgcc aaaaaatgaa aactgtggac aagctctgtc cgtttgctgg
     6601 caactgcaag ggtctcaatc ctatttgtaa ttattgaata ataaaacaat tataaatgct
     6661 aaatttgttt tttattaacg atacaaacca aacgcaacaa gaacatttgt agtattatct
     6721 ataattgaaa acgcgtagtt ataatcgctg aggtaatatt taaaatcatt ttcaaatgat
     6781 tcacagttaa tttgcgacaa tataatttta ttttcacata aactagacgc cttgtcgtct
     6841 tcttcttcgt attccttctc tttttcattt ttctcctcat aaaaattaac atagttatta
     6901 tcgtatccat atatgtatct atcgtataga gtaaattttt tgttgtcata aatatatatg
     6961 tcttttttaa tggggtgtat agtaccgctg cgcatagttt ttctgtaatt tacaacagtg
     7021 ctattttctg gtagttcttc ggagtgtgtt gctttaatta ttaaatttat ataatcaatg
     7081 aatttgggat cgtcggtttt gtacaatatg ttgccggcat agtacgcagc ttcttctagt
     7141 tcaattacac cattttttag cagcaccgga ttaacataac tttccaaaat gttgtacgaa
     7201 ccgttaaaca aaaacagttc acctcccttt tctatactat tgtctgcgag cagttgtttg
     7261 ttgttaaaaa taacagccat tgtaatgaga cgcacaaact aatatcaca
//
