LOCUS       sequence-232363-        7383 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal MBP, cleavable with 3C and 
            monomeric GFP and C-terminal TwinStrep, cleavable with 3C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7383)
  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 7383)
  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: pOCC193
FEATURES             Location/Qualifiers
     source          1..7383
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..92
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     17..34
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     regulatory      134..143
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             146..1246
                     /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     1220..1243
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1301..1324
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(1325..1327,1328..1330,1331..2041)
                     /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(1370..1391)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1631..1650)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1978..1999
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2180..2485
                     /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     2364..2383
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2523..2546
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(2547..2552,2553..2576,2577..2612,2613..2636)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     polyA_signal    2772..2906
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2809..2828)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2863..2882
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2979..3351
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3703..3720)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3813..4401)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3893..3912)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4572..5363,5364..5432))
                     /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     5195..5214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5433..5537)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5838..5860
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5852..5869
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5853..5869
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5880..5899
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5880..5898
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6509..6633
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6671..7276)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
ORIGIN
        1 atcatggaga taattaaaat gataaccatc tcgcaaataa ataagtattt tactgttttc
       61 gtaacagttt tgtaataaaa aaacctataa atattccgga ttattcatac cgtcccacca
      121 tcgggcgcgg atcgccacca tgggcatgaa aatcgaagaa ggtaaactgg taatctggat
      181 taacggcgat aaaggctata acggtctcgc tgaagtcggt aagaaattcg agaaagatac
      241 cggaattaaa gtcaccgttg agcatccgga taaactggaa gagaaattcc cacaggttgc
      301 ggcaactggc gatggccctg acattatctt ctgggcacac gaccgctttg gtggctacgc
      361 tcaatctggc ctgttggctg aaatcacccc ggacaaagcg ttccaggaca agctgtatcc
      421 gtttacctgg gatgccgtac gttacaacgg caagctgatt gcttacccga tcgctgttga
      481 agcgttatcg ctgatttata acaaagatct gctgccgaac ccgccaaaaa cctgggaaga
      541 gatcccggcg ctggataaag aactgaaagc gaaaggtaag agcgcgctga tgttcaacct
      601 gcaagaaccg tacttcacct ggccgctgat tgctgctgac gggggttatg cgttcaagta
      661 tgaaaacggc aagtacgaca ttaaagacgt gggcgtggat aacgctggcg cgaaagcggg
      721 tctgaccttc ctggttgacc tgattaaaaa caaacacatg aatgcagaca ccgattactc
      781 catcgcagaa gctgccttta ataaaggcga aacagcgatg accatcaacg gcccgtgggc
      841 atggtccaac atcgacacca gcaaagtgaa ttatggtgta acggtactgc cgaccttcaa
      901 gggtcaacca tccaaaccgt tcgttggcgt gctgagcgca ggtattaacg ccgccagtcc
      961 gaacaaagag ctggcaaaag agttcctcga aaactatctg ctgactgatg aaggtctgga
     1021 agcggttaat aaagacaaac cgctgggtgc cgtagcgctg aagtcttacg aggaagagtt
     1081 ggtgaaagat ccgcgtattg ccgccactat ggaaaacgcc cagaaaggtg aaatcatgcc
     1141 gaacatcccg cagatgtccg ctttctggta tgccgtgcgt actgcggtga tcaacgccgc
     1201 cagcggtcgt cagactgtcg atgaagccct gaaagacgcg cagactaatt cgagctcgaa
     1261 caacaacaac aataacaata acaacaacag cagcggcaga ctggaagttc tgttccaggg
     1321 gccgatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct
     1381 ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac
     1441 ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc
     1501 caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat
     1561 gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat
     1621 cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac
     1681 cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg
     1741 gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa
     1801 gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct
     1861 cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa
     1921 ccactacctg agcacccagt ccaaactgag caaagacccc aacgagaagc gcgatcacat
     1981 ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa
     2041 gggatccgct ggctccgctg ctggttctgg cgcggccgca agcgctatgg ccacaacctg
     2101 ttacattgca caagataaaa atatatcatc acgaacagta aaactgtctg cttacataaa
     2161 cagtaataca aggggtgtta tgcagtttaa ggtttacacc tataaaagag agagccgtta
     2221 tcgtctgttt gtggatgtac agagtgatat tattgacacg cccgggcgac ggatggtgat
     2281 ccccctggcc agtgcccgtc tgctgtcaga taaagtctcc cgtgaacttt acccggtggt
     2341 gcatatcggg gatgaaagct ggcgcatgat gaccaccgat atggccagtg tgccggtctc
     2401 cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca aaaacgccat
     2461 taacctgatg ttctggggaa tataaatgtc aggctccctt atacacagcc agtggcgcgc
     2521 cgctggaagt tctgttccag gggccgagcg cttggagcca tccgcagttt gaaaaaggtg
     2581 gtggtagcgg tggtggttca ggtggtagtg catggtcaca ccctcagttt gagaaataat
     2641 agaagcttgt cgagaagtac tagaggatca taatcagcca taccacattt gtagaggttt
     2701 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     2761 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     2821 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     2881 tcaatgtatc ttatcatgtc tggatctgat cactgcttga gcctaggaga tccgaaccag
     2941 ataagtgaaa gcctgcatcc gcagatctta attaataaaa cacgatacat tgttattagt
     3001 acatttatta agcgctagat tctgtgcgtt gttgatttac agacaattgt tgtacgtatt
     3061 ttaataattc attaaattta taatctttag ggtggtatgt tagagcgaaa atcaaatgat
     3121 tttcagcgtc tttatatctg aatttaaata ttaaatcctc aatagatttg taaaataggt
     3181 ttcgattagt ttcaaacaag ggttgttttt ccgaaccgat ggctggacta tctaatggat
     3241 tttcgctcaa cgccacaaaa cttgccaaat cttgtagcag caatctagct ttgtcgatat
     3301 tcgtttgtgt tttgttttgt aataaaggtt cgacgtcgtt caaaatatta tgcgcttttg
     3361 tatttctttc atcactgtcg ttagtgtaca attgactcga cgtaaacacg ttaaataacg
     3421 cttggacata tttaacatcg ggcgtgttag ctttattagg ccgattatcg tcgtcgtccc
     3481 aaccctcgtc gttagaagtt gcttccgaag acgattttgc catagccaca cgacgcctat
     3541 taattgtgtc ggctaacacg tccgcgatca aatttgtagt tgagcttttt gggcctcatg
     3601 ggccttccgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaaca
     3661 tggtcatagc tgtttccttg cgtattgggc gctctccgct tcctcgctca ctgactcgct
     3721 gcgctcggtc gttcgggtaa agcctggggt gcctaatgag caaaaggcca gcaaaaggcc
     3781 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     3841 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3901 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3961 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     4021 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     4081 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     4141 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     4201 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     4261 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     4321 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     4381 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     4441 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     4501 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     4561 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     4621 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     4681 ccatctggcc ccagtgctgc aatgataccg cgagaaccac gctcaccggc tccagattta
     4741 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     4801 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     4861 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     4921 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4981 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     5041 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     5101 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     5161 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     5221 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     5281 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     5341 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     5401 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     5461 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     5521 caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata
     5581 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     5641 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtggccgct
     5701 acagggcgct cccattcgcc attcaggctg cgcaactgtt gggaagggcg tttcggtgcg
     5761 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     5821 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgacgt
     5881 aatacgactc actatagggc gaattggcgg aaggccgtca aggccgtgac gttaaaacta
     5941 ttaagccatc caatcgaccg ttagtcgaat caggaccgct ggtgcgagaa gccgcgaagt
     6001 atggcgaatg catcgtataa cgtgtggagt ccgctcatta gagcgtcatg tttagacaag
     6061 aaagctacat atttaattga tcccgatgat tttattgata aattgaccct aactccatac
     6121 acggtattct acaatggcgg ggttttggtc aaaatttccg gactgcgatt gtacatgctg
     6181 ttaacggctc cgcccactat taatgaaatt aaaaattcca attttaaaaa acgcagcaag
     6241 agaaacattt gtatgaaaga atgcgtagaa ggaaagaaaa atgtcgtgga catgctgaac
     6301 aacaagatta atatgcctcc gtgtataaaa aaaatattga acgatttgaa agaaaacaat
     6361 gtaccgcgcg gcggtatgta caggaagagg tttatactaa actgttacat tgcaaacgtg
     6421 gtttcgtgtg ccaagtgtga aaaccgatgt ttaatcaagg ctctgacgca tttctacaac
     6481 cacgactcca agtgtgtggg tgaagtcatg catcttttaa tcaaatccca agatgtgtat
     6541 aaaccaccaa actgccaaaa aatgaaaact gtggacaagc tctgtccgtt tgctggcaac
     6601 tgcaagggtc tcaatcctat ttgtaattat tgaataataa aacaattata aatgctaaat
     6661 ttgtttttta ttaacgatac aaaccaaacg caacaagaac atttgtagta ttatctataa
     6721 ttgaaaacgc gtagttataa tcgctgaggt aatatttaaa atcattttca aatgattcac
     6781 agttaatttg cgacaatata attttatttt cacataaact agacgccttg tcgtcttctt
     6841 cttcgtattc cttctctttt tcatttttct cctcataaaa attaacatag ttattatcgt
     6901 atccatatat gtatctatcg tatagagtaa attttttgtt gtcataaata tatatgtctt
     6961 ttttaatggg gtgtatagta ccgctgcgca tagtttttct gtaatttaca acagtgctat
     7021 tttctggtag ttcttcggag tgtgttgctt taattattaa atttatataa tcaatgaatt
     7081 tgggatcgtc ggttttgtac aatatgttgc cggcatagta cgcagcttct tctagttcaa
     7141 ttacaccatt ttttagcagc accggattaa cataactttc caaaatgttg tacgaaccgt
     7201 taaacaaaaa cagttcacct cccttttcta tactattgtc tgcgagcagt tgtttgttgt
     7261 taaaaataac agccattgta atgagacgca caaactaata tcacaagctg ccaccatttt
     7321 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     7381 tag
//
