LOCUS       sequence-232364-        6681 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for rec. protein with N-terminal TwinStrep-MBP tag, cleavable with 
            3C and C-terminal HIS6, cleavable with 3C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6681)
  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 6681)
  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: pOCC305
FEATURES             Location/Qualifiers
     source          1..6681
                     /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             join(146..151,152..175,176..211,212..235)
                     /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"
     CDS             248..1348
                     /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     1322..1345
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1403..1426
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1535..1840
                     /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     1719..1738
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1878..1901
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1911..1928
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2070..2204
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2107..2126)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2161..2180
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2277..2649
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3001..3018)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3111..3699)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3191..3210)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3870..4661,4662..4730))
                     /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     4493..4512
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4731..4835)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5136..5158
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5150..5167
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5151..5167
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5178..5197
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5178..5196
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5807..5931
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5969..6574)
                     /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 tgggcagcgc ttggagccat ccgcagtttg aaaaaggtgg
      181 tggtagcggt ggtggttcag gtggtagtgc atggtcacac cctcagtttg agaaaagcag
      241 cggcagaatg aaaatcgaag aaggtaaact ggtaatctgg attaacggcg ataaaggcta
      301 taacggtctc gctgaagtcg gtaagaaatt cgagaaagat accggaatta aagtcaccgt
      361 tgagcatccg gataaactgg aagagaaatt cccacaggtt gcggcaactg gcgatggccc
      421 tgacattatc ttctgggcac acgaccgctt tggtggctac gctcaatctg gcctgttggc
      481 tgaaatcacc ccggacaaag cgttccagga caagctgtat ccgtttacct gggatgccgt
      541 acgttacaac ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat cgctgattta
      601 taacaaagat ctgctgccga acccgccaaa aacctgggaa gagatcccgg cgctggataa
      661 agaactgaaa gcgaaaggta agagcgcgct gatgttcaac ctgcaagaac cgtacttcac
      721 ctggccgctg attgctgctg acgggggtta tgcgttcaag tatgaaaacg gcaagtacga
      781 cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg ggtctgacct tcctggttga
      841 cctgattaaa aacaaacaca tgaatgcaga caccgattac tccatcgcag aagctgcctt
      901 taataaaggc gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca acatcgacac
      961 cagcaaagtg aattatggtg taacggtact gccgaccttc aagggtcaac catccaaacc
     1021 gttcgttggc gtgctgagcg caggtattaa cgccgccagt ccgaacaaag agctggcaaa
     1081 agagttcctc gaaaactatc tgctgactga tgaaggtctg gaagcggtta ataaagacaa
     1141 accgctgggt gccgtagcgc tgaagtctta cgaggaagag ttggtgaaag atccgcgtat
     1201 tgccgccact atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc cgcagatgtc
     1261 cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc gtcagactgt
     1321 cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg aacaacaaca acaataacaa
     1381 taacaacaac agcagcggca gactggaagt tctgttccag gggccggcgg ccgcaagcgc
     1441 tatggccaca acctgttaca ttgcacaaga taaaaatata tcatcacgaa cagtaaaact
     1501 gtctgcttac ataaacagta atacaagggg tgttatgcag tttaaggttt acacctataa
     1561 aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg
     1621 gcgacggatg gtgatccccc tggccagtgc ccgtctgctg tcagataaag tctcccgtga
     1681 actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc
     1741 cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga
     1801 catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggct cccttataca
     1861 cagccagtgg cgcgccgctg gaagttctgt tccaggggcc gggcagcagc catcaccatc
     1921 accatcacag tggataatag aagcttgtcg agaagtacta gaggatcata atcagccata
     1981 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
     2041 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
     2101 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
     2161 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatctgatca ctgcttgagc
     2221 ctaggagatc cgaaccagat aagtgaaagc ctgcatccgc agatcttaat taataaaaca
     2281 cgatacattg ttattagtac atttattaag cgctagattc tgtgcgttgt tgatttacag
     2341 acaattgttg tacgtatttt aataattcat taaatttata atctttaggg tggtatgtta
     2401 gagcgaaaat caaatgattt tcagcgtctt tatatctgaa tttaaatatt aaatcctcaa
     2461 tagatttgta aaataggttt cgattagttt caaacaaggg ttgtttttcc gaaccgatgg
     2521 ctggactatc taatggattt tcgctcaacg ccacaaaact tgccaaatct tgtagcagca
     2581 atctagcttt gtcgatattc gtttgtgttt tgttttgtaa taaaggttcg acgtcgttca
     2641 aaatattatg cgcttttgta tttctttcat cactgtcgtt agtgtacaat tgactcgacg
     2701 taaacacgtt aaataacgct tggacatatt taacatcggg cgtgttagct ttattaggcc
     2761 gattatcgtc gtcgtcccaa ccctcgtcgt tagaagttgc ttccgaagac gattttgcca
     2821 tagccacacg acgcctatta attgtgtcgg ctaacacgtc cgcgatcaaa tttgtagttg
     2881 agctttttgg gcctcatggg ccttccgctc actgcccgct ttccagtcgg gaaacctgtc
     2941 gtgccagctg cattaacatg gtcatagctg tttccttgcg tattgggcgc tctccgcttc
     3001 ctcgctcact gactcgctgc gctcggtcgt tcgggtaaag cctggggtgc ctaatgagca
     3061 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3121 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3181 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3241 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3301 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3361 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3421 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3481 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3541 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3601 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3661 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3721 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3781 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3841 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3901 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3961 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agaaccacgc
     4021 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4081 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4141 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4201 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4261 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4321 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4381 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4441 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4501 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4561 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4621 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4681 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4741 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4801 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4861 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     4921 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     4981 tagggttgag tggccgctac agggcgctcc cattcgccat tcaggctgcg caactgttgg
     5041 gaagggcgtt tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     5101 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     5161 ggccagtgag cgcgacgtaa tacgactcac tatagggcga attggcggaa ggccgtcaag
     5221 gccgtgacgt taaaactatt aagccatcca atcgaccgtt agtcgaatca ggaccgctgg
     5281 tgcgagaagc cgcgaagtat ggcgaatgca tcgtataacg tgtggagtcc gctcattaga
     5341 gcgtcatgtt tagacaagaa agctacatat ttaattgatc ccgatgattt tattgataaa
     5401 ttgaccctaa ctccatacac ggtattctac aatggcgggg ttttggtcaa aatttccgga
     5461 ctgcgattgt acatgctgtt aacggctccg cccactatta atgaaattaa aaattccaat
     5521 tttaaaaaac gcagcaagag aaacatttgt atgaaagaat gcgtagaagg aaagaaaaat
     5581 gtcgtggaca tgctgaacaa caagattaat atgcctccgt gtataaaaaa aatattgaac
     5641 gatttgaaag aaaacaatgt accgcgcggc ggtatgtaca ggaagaggtt tatactaaac
     5701 tgttacattg caaacgtggt ttcgtgtgcc aagtgtgaaa accgatgttt aatcaaggct
     5761 ctgacgcatt tctacaacca cgactccaag tgtgtgggtg aagtcatgca tcttttaatc
     5821 aaatcccaag atgtgtataa accaccaaac tgccaaaaaa tgaaaactgt ggacaagctc
     5881 tgtccgtttg ctggcaactg caagggtctc aatcctattt gtaattattg aataataaaa
     5941 caattataaa tgctaaattt gttttttatt aacgatacaa accaaacgca acaagaacat
     6001 ttgtagtatt atctataatt gaaaacgcgt agttataatc gctgaggtaa tatttaaaat
     6061 cattttcaaa tgattcacag ttaatttgcg acaatataat tttattttca cataaactag
     6121 acgccttgtc gtcttcttct tcgtattcct tctctttttc atttttctcc tcataaaaat
     6181 taacatagtt attatcgtat ccatatatgt atctatcgta tagagtaaat tttttgttgt
     6241 cataaatata tatgtctttt ttaatggggt gtatagtacc gctgcgcata gtttttctgt
     6301 aatttacaac agtgctattt tctggtagtt cttcggagtg tgttgcttta attattaaat
     6361 ttatataatc aatgaatttg ggatcgtcgg ttttgtacaa tatgttgccg gcatagtacg
     6421 cagcttcttc tagttcaatt acaccatttt ttagcagcac cggattaaca taactttcca
     6481 aaatgttgta cgaaccgtta aacaaaaaca gttcacctcc cttttctata ctattgtctg
     6541 cgagcagttg tttgttgtta aaaataacag ccattgtaat gagacgcaca aactaatatc
     6601 acaagctgcc accattttgt tcgcccagga ctctagctat agttctagtg gttggctacg
     6661 tatactccgg aatattaata g
//
