LOCUS       sequence-224685-        7318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for rec. protein with N-terminal MBP tag, cleavable with 3C and 
            C-terminal mCherry, cleavable with TEV and HIS6, cleavable with 3C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7318)
  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 7318)
  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: pOCC240
FEATURES             Location/Qualifiers
     source          1..7318
                     /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..1324
                     /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     1298..1321
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1379..1402
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1511..1816
                     /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     1695..1714
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1863..1883
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1884..2591
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2029..2047)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2297..2316
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2592..2615
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2625..2642
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2784..2918
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2821..2840)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2875..2894
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2991..3363
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3715..3732)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3825..4413)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3905..3924)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4584..5375,5376..5444))
                     /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     5207..5226
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5446..5550)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5851..5873
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5865..5882
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5866..5882
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5893..5912
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5893..5911
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6522..6646
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6684..7289)
                     /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 ggcatgaaaa tcgaagaagg
      241 taaactggta atctggatta acggcgataa aggctataac ggtctcgctg aagtcggtaa
      301 gaaattcgag aaagataccg gaattaaagt caccgttgag catccggata aactggaaga
      361 gaaattccca caggttgcgg caactggcga tggccctgac attatcttct gggcacacga
      421 ccgctttggt ggctacgctc aatctggcct gttggctgaa atcaccccgg acaaagcgtt
      481 ccaggacaag ctgtatccgt ttacctggga tgccgtacgt tacaacggca agctgattgc
      541 ttacccgatc gctgttgaag cgttatcgct gatttataac aaagatctgc tgccgaaccc
      601 gccaaaaacc tgggaagaga tcccggcgct ggataaagaa ctgaaagcga aaggtaagag
      661 cgcgctgatg ttcaacctgc aagaaccgta cttcacctgg ccgctgattg ctgctgacgg
      721 gggttatgcg ttcaagtatg aaaacggcaa gtacgacatt aaagacgtgg gcgtggataa
      781 cgctggcgcg aaagcgggtc tgaccttcct ggttgacctg attaaaaaca aacacatgaa
      841 tgcagacacc gattactcca tcgcagaagc tgcctttaat aaaggcgaaa cagcgatgac
      901 catcaacggc ccgtgggcat ggtccaacat cgacaccagc aaagtgaatt atggtgtaac
      961 ggtactgccg accttcaagg gtcaaccatc caaaccgttc gttggcgtgc tgagcgcagg
     1021 tattaacgcc gccagtccga acaaagagct ggcaaaagag ttcctcgaaa actatctgct
     1081 gactgatgaa ggtctggaag cggttaataa agacaaaccg ctgggtgccg tagcgctgaa
     1141 gtcttacgag gaagagttgg tgaaagatcc gcgtattgcc gccactatgg aaaacgccca
     1201 gaaaggtgaa atcatgccga acatcccgca gatgtccgct ttctggtatg ccgtgcgtac
     1261 tgcggtgatc aacgccgcca gcggtcgtca gactgtcgat gaagccctga aagacgcgca
     1321 gactaattcg agctcgaaca acaacaacaa taacaataac aacaacagca gcggcagact
     1381 ggaagttctg ttccaggggc cggcggccgc aagcgctatg gccacaacct gttacattgc
     1441 acaagataaa aatatatcat cacgaacagt aaaactgtct gcttacataa acagtaatac
     1501 aaggggtgtt atgcagttta aggtttacac ctataaaaga gagagccgtt atcgtctgtt
     1561 tgtggatgta cagagtgata ttattgacac gcccgggcga cggatggtga tccccctggc
     1621 cagtgcccgt ctgctgtcag ataaagtctc ccgtgaactt tacccggtgg tgcatatcgg
     1681 ggatgaaagc tggcgcatga tgaccaccga tatggccagt gtgccggtct ccgttatcgg
     1741 ggaagaagtg gctgatctca gccaccgcga aaatgacatc aaaaacgcca ttaacctgat
     1801 gttctgggga atataaatgt caggctccct tatacacagc cagtggcgcg ccgggatcca
     1861 gcgaaaactt gtatttccag ggcatggtga gcaagggcga ggaggataac atggccatca
     1921 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
     1981 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     2041 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
     2101 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
     2161 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga
     2221 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
     2281 acttcccctc cgacggcccc gtaatgcaga agaagacgat gggctgggag gcctcctccg
     2341 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     2401 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     2461 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     2521 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     2581 agctgtacaa gctggaagtt ctgttccagg ggccgggcag cagccatcac catcaccatc
     2641 acagtggata atagaagctt gtcgagaagt actagaggat cataatcagc cataccacat
     2701 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     2761 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     2821 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     2881 tgtccaaact catcaatgta tcttatcatg tctggatctg atcactgctt gagcctagga
     2941 gatccgaacc agataagtga aagcctgcat ccgcagatct taattaataa aacacgatac
     3001 attgttatta gtacatttat taagcgctag attctgtgcg ttgttgattt acagacaatt
     3061 gttgtacgta ttttaataat tcattaaatt tataatcttt agggtggtat gttagagcga
     3121 aaatcaaatg attttcagcg tctttatatc tgaatttaaa tattaaatcc tcaatagatt
     3181 tgtaaaatag gtttcgatta gtttcaaaca agggttgttt ttccgaaccg atggctggac
     3241 tatctaatgg attttcgctc aacgccacaa aacttgccaa atcttgtagc agcaatctag
     3301 ctttgtcgat attcgtttgt gttttgtttt gtaataaagg ttcgacgtcg ttcaaaatat
     3361 tatgcgcttt tgtatttctt tcatcactgt cgttagtgta caattgactc gacgtaaaca
     3421 cgttaaataa cgcttggaca tatttaacat cgggcgtgtt agctttatta ggccgattat
     3481 cgtcgtcgtc ccaaccctcg tcgttagaag ttgcttccga agacgatttt gccatagcca
     3541 cacgacgcct attaattgtg tcggctaaca cgtccgcgat caaatttgta gttgagcttt
     3601 ttgggcctca tgggccttcc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     3661 gctgcattaa catggtcata gctgtttcct tgcgtattgg gcgctctccg cttcctcgct
     3721 cactgactcg ctgcgctcgg tcgttcgggt aaagcctggg gtgcctaatg agcaaaaggc
     3781 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     3841 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     3901 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     3961 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4021 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4081 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4141 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4201 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4261 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4321 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4381 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4441 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4501 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     4561 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     4621 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     4681 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     4741 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     4801 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     4861 tcgccagtta atagtttgcg caacgttgtt gccattgctg caggcatcgt ggtgtcacgc
     4921 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     4981 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5041 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5101 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5161 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa cacgggataa taccgcgcca
     5221 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5281 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5341 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5401 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcattactct tcctttttca
     5461 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     5521 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctaaatt
     5581 gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt
     5641 aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg
     5701 ttgagtggcc gctacagggc gctcccattc gccattcagg ctgcgcaact gttgggaagg
     5761 gcgtttcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     5821 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     5881 gtgagcgcga cgtaatacga ctcactatag ggcgaattgg cggaaggccg tcaaggccgt
     5941 gacgttaaaa ctattaagcc atccaatcga ccgttagtcg aatcaggacc gctggtgcga
     6001 gaagccgcga agtatggcga atgcatcgta taacgtgtgg agtccgctca ttagagcgtc
     6061 atgtttagac aagaaagcta catatttaat tgatcccgat gattttattg ataaattgac
     6121 cctaactcca tacacggtat tctacaatgg cggggttttg gtcaaaattt ccggactgcg
     6181 attgtacatg ctgttaacgg ctccgcccac tattaatgaa attaaaaatt ccaattttaa
     6241 aaaacgcagc aagagaaaca tttgtatgaa agaatgcgta gaaggaaaga aaaatgtcgt
     6301 ggacatgctg aacaacaaga ttaatatgcc tccgtgtata aaaaaaatat tgaacgattt
     6361 gaaagaaaac aatgtaccgc gcggcggtat gtacaggaag aggtttatac taaactgtta
     6421 cattgcaaac gtggtttcgt gtgccaagtg tgaaaaccga tgtttaatca aggctctgac
     6481 gcatttctac aaccacgact ccaagtgtgt gggtgaagtc atgcatcttt taatcaaatc
     6541 ccaagatgtg tataaaccac caaactgcca aaaaatgaaa actgtggaca agctctgtcc
     6601 gtttgctggc aactgcaagg gtctcaatcc tatttgtaat tattgaataa taaaacaatt
     6661 ataaatgcta aatttgtttt ttattaacga tacaaaccaa acgcaacaag aacatttgta
     6721 gtattatcta taattgaaaa cgcgtagtta taatcgctga ggtaatattt aaaatcattt
     6781 tcaaatgatt cacagttaat ttgcgacaat ataattttat tttcacataa actagacgcc
     6841 ttgtcgtctt cttcttcgta ttccttctct ttttcatttt tctcctcata aaaattaaca
     6901 tagttattat cgtatccata tatgtatcta tcgtatagag taaatttttt gttgtcataa
     6961 atatatatgt cttttttaat ggggtgtata gtaccgctgc gcatagtttt tctgtaattt
     7021 acaacagtgc tattttctgg tagttcttcg gagtgtgttg ctttaattat taaatttata
     7081 taatcaatga atttgggatc gtcggttttg tacaatatgt tgccggcata gtacgcagct
     7141 tcttctagtt caattacacc attttttagc agcaccggat taacataact ttccaaaatg
     7201 ttgtacgaac cgttaaacaa aaacagttca cctccctttt ctatactatt gtctgcgagc
     7261 agttgtttgt tgttaaaaat aacagccatt gtaatgagac gcacaaacta atatcaca
//
