LOCUS       sequence-232282-        7143 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 N-terminal SNAP, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7143)
  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 7143)
  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: pOCC176
FEATURES             Location/Qualifiers
     source          1..7143
                     /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             152..169
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             182..1282
                     /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     1256..1279
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1337..1360
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1361..1906
                     /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"
     CDS             1925..1945
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2054..2359
                     /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     2238..2257
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    2532..2666
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2569..2588)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2623..2642
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2739..3111
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3463..3480)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3573..4161)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3653..3672)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4332..5123,5124..5192))
                     /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     4955..4974
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5193..5297)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5598..5620
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5612..5629
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5613..5629
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5640..5659
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5640..5658
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6269..6393
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6431..7036)
                     /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 tgggcagcag ccatcaccat caccatcaca gcagcggcag
      181 aatgaaaatc gaagaaggta aactggtaat ctggattaac ggcgataaag gctataacgg
      241 tctcgctgaa gtcggtaaga aattcgagaa agataccgga attaaagtca ccgttgagca
      301 tccggataaa ctggaagaga aattcccaca ggttgcggca actggcgatg gccctgacat
      361 tatcttctgg gcacacgacc gctttggtgg ctacgctcaa tctggcctgt tggctgaaat
      421 caccccggac aaagcgttcc aggacaagct gtatccgttt acctgggatg ccgtacgtta
      481 caacggcaag ctgattgctt acccgatcgc tgttgaagcg ttatcgctga tttataacaa
      541 agatctgctg ccgaacccgc caaaaacctg ggaagagatc ccggcgctgg ataaagaact
      601 gaaagcgaaa ggtaagagcg cgctgatgtt caacctgcaa gaaccgtact tcacctggcc
      661 gctgattgct gctgacgggg gttatgcgtt caagtatgaa aacggcaagt acgacattaa
      721 agacgtgggc gtggataacg ctggcgcgaa agcgggtctg accttcctgg ttgacctgat
      781 taaaaacaaa cacatgaatg cagacaccga ttactccatc gcagaagctg cctttaataa
      841 aggcgaaaca gcgatgacca tcaacggccc gtgggcatgg tccaacatcg acaccagcaa
      901 agtgaattat ggtgtaacgg tactgccgac cttcaagggt caaccatcca aaccgttcgt
      961 tggcgtgctg agcgcaggta ttaacgccgc cagtccgaac aaagagctgg caaaagagtt
     1021 cctcgaaaac tatctgctga ctgatgaagg tctggaagcg gttaataaag acaaaccgct
     1081 gggtgccgta gcgctgaagt cttacgagga agagttggtg aaagatccgc gtattgccgc
     1141 cactatggaa aacgcccaga aaggtgaaat catgccgaac atcccgcaga tgtccgcttt
     1201 ctggtatgcc gtgcgtactg cggtgatcaa cgccgccagc ggtcgtcaga ctgtcgatga
     1261 agccctgaaa gacgcgcaga ctaattcgag ctcgaacaac aacaacaata acaataacaa
     1321 caacagcagc ggcagactgg aagttctgtt ccaggggccg atggacaaag actgcgaaat
     1381 gaagcgcacc accctggata gccctctggg caagctggaa ctgtctgggt gcgaacaggg
     1441 cctgcaccgt atcatcttcc tgggcaaagg aacatctgcc gccgacgccg tggaagtgcc
     1501 tgccccagcc gccgtgctgg gcggaccaga gccactgatg caggccaccg cctggctcaa
     1561 cgcctacttt caccagcctg aggccatcga ggagttccct gtgccagccc tgcaccaccc
     1621 agtgttccag caggagagct ttacccgcca ggtgctgtgg aaactgctga aagtggtgaa
     1681 gttcggagag gtcatcagct acagccacct ggccgccctg gccggcaatc ccgccgccac
     1741 cgccgccgtg aaaaccgccc tgagcggaaa tcccgtgccc attctgatcc cctgccaccg
     1801 ggtggtgcag ggcgacctgg acgtgggggg ctacgagggc gggctcgccg tgaaagagtg
     1861 gctgctggcc cacgagggcc acagactggg caagcctggg ctgggtggat ccagcagcgg
     1921 cagagaaaac ttgtatttcc agggcgcggc cgcaagcgct atggccacaa cctgttacat
     1981 tgcacaagat aaaaatatat catcacgaac agtaaaactg tctgcttaca taaacagtaa
     2041 tacaaggggt gttatgcagt ttaaggttta cacctataaa agagagagcc gttatcgtct
     2101 gtttgtggat gtacagagtg atattattga cacgcccggg cgacggatgg tgatccccct
     2161 ggccagtgcc cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg tggtgcatat
     2221 cggggatgaa agctggcgca tgatgaccac cgatatggcc agtgtgccgg tctccgttat
     2281 cggggaagaa gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg ccattaacct
     2341 gatgttctgg ggaatataaa tgtcaggctc ccttatacac agccagtggc gcgccgtaat
     2401 agaagcttgt cgagaagtac tagaggatca taatcagcca taccacattt gtagaggttt
     2461 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     2521 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     2581 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     2641 tcaatgtatc ttatcatgtc tggatctgat cactgcttga gcctaggaga tccgaaccag
     2701 ataagtgaaa gcctgcatcc gcagatctta attaataaaa cacgatacat tgttattagt
     2761 acatttatta agcgctagat tctgtgcgtt gttgatttac agacaattgt tgtacgtatt
     2821 ttaataattc attaaattta taatctttag ggtggtatgt tagagcgaaa atcaaatgat
     2881 tttcagcgtc tttatatctg aatttaaata ttaaatcctc aatagatttg taaaataggt
     2941 ttcgattagt ttcaaacaag ggttgttttt ccgaaccgat ggctggacta tctaatggat
     3001 tttcgctcaa cgccacaaaa cttgccaaat cttgtagcag caatctagct ttgtcgatat
     3061 tcgtttgtgt tttgttttgt aataaaggtt cgacgtcgtt caaaatatta tgcgcttttg
     3121 tatttctttc atcactgtcg ttagtgtaca attgactcga cgtaaacacg ttaaataacg
     3181 cttggacata tttaacatcg ggcgtgttag ctttattagg ccgattatcg tcgtcgtccc
     3241 aaccctcgtc gttagaagtt gcttccgaag acgattttgc catagccaca cgacgcctat
     3301 taattgtgtc ggctaacacg tccgcgatca aatttgtagt tgagcttttt gggcctcatg
     3361 ggccttccgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaaca
     3421 tggtcatagc tgtttccttg cgtattgggc gctctccgct tcctcgctca ctgactcgct
     3481 gcgctcggtc gttcgggtaa agcctggggt gcctaatgag caaaaggcca gcaaaaggcc
     3541 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     3601 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3661 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3721 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3781 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3841 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3901 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3961 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     4021 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     4081 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     4141 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     4201 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     4261 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     4321 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     4381 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     4441 ccatctggcc ccagtgctgc aatgataccg cgagaaccac gctcaccggc tccagattta
     4501 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     4561 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     4621 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     4681 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4741 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4801 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4861 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4921 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4981 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     5041 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     5101 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     5161 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     5221 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     5281 caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata
     5341 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     5401 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtggccgct
     5461 acagggcgct cccattcgcc attcaggctg cgcaactgtt gggaagggcg tttcggtgcg
     5521 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     5581 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgacgt
     5641 aatacgactc actatagggc gaattggcgg aaggccgtca aggccgtgac gttaaaacta
     5701 ttaagccatc caatcgaccg ttagtcgaat caggaccgct ggtgcgagaa gccgcgaagt
     5761 atggcgaatg catcgtataa cgtgtggagt ccgctcatta gagcgtcatg tttagacaag
     5821 aaagctacat atttaattga tcccgatgat tttattgata aattgaccct aactccatac
     5881 acggtattct acaatggcgg ggttttggtc aaaatttccg gactgcgatt gtacatgctg
     5941 ttaacggctc cgcccactat taatgaaatt aaaaattcca attttaaaaa acgcagcaag
     6001 agaaacattt gtatgaaaga atgcgtagaa ggaaagaaaa atgtcgtgga catgctgaac
     6061 aacaagatta atatgcctcc gtgtataaaa aaaatattga acgatttgaa agaaaacaat
     6121 gtaccgcgcg gcggtatgta caggaagagg tttatactaa actgttacat tgcaaacgtg
     6181 gtttcgtgtg ccaagtgtga aaaccgatgt ttaatcaagg ctctgacgca tttctacaac
     6241 cacgactcca agtgtgtggg tgaagtcatg catcttttaa tcaaatccca agatgtgtat
     6301 aaaccaccaa actgccaaaa aatgaaaact gtggacaagc tctgtccgtt tgctggcaac
     6361 tgcaagggtc tcaatcctat ttgtaattat tgaataataa aacaattata aatgctaaat
     6421 ttgtttttta ttaacgatac aaaccaaacg caacaagaac atttgtagta ttatctataa
     6481 ttgaaaacgc gtagttataa tcgctgaggt aatatttaaa atcattttca aatgattcac
     6541 agttaatttg cgacaatata attttatttt cacataaact agacgccttg tcgtcttctt
     6601 cttcgtattc cttctctttt tcatttttct cctcataaaa attaacatag ttattatcgt
     6661 atccatatat gtatctatcg tatagagtaa attttttgtt gtcataaata tatatgtctt
     6721 ttttaatggg gtgtatagta ccgctgcgca tagtttttct gtaatttaca acagtgctat
     6781 tttctggtag ttcttcggag tgtgttgctt taattattaa atttatataa tcaatgaatt
     6841 tgggatcgtc ggttttgtac aatatgttgc cggcatagta cgcagcttct tctagttcaa
     6901 ttacaccatt ttttagcagc accggattaa cataactttc caaaatgttg tacgaaccgt
     6961 taaacaaaaa cagttcacct cccttttcta tactattgtc tgcgagcagt tgtttgttgt
     7021 taaaaataac agccattgta atgagacgca caaactaata tcacaagctg ccaccatttt
     7081 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     7141 tag
//
