LOCUS       sequence-224677-        7480 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 HALO tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7480)
  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 7480)
  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: pOCC267
FEATURES             Location/Qualifiers
     source          1..7480
                     /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             230..247
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             260..1360
                     /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     1334..1357
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1415..1438
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1439..2329
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     CDS             2468..2773
                     /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     2652..2671
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    2946..3080
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2983..3002)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3037..3056
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           3153..3525
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3877..3894)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3987..4575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4067..4086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4746..5537,5538..5606))
                     /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     5369..5388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5608..5712)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6013..6035
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6027..6044
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6028..6044
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6055..6074
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6055..6073
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6684..6808
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6846..7451)
                     /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 ggcagcagcc atcaccatca
      241 ccatcacagc agcggcagaa tgaaaatcga agaaggtaaa ctggtaatct ggattaacgg
      301 cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat
      361 taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac
      421 tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc
      481 tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac
      541 ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt
      601 atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc
      661 ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga
      721 accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa
      781 cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac
      841 cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc
      901 agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc
      961 caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca
     1021 accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa
     1081 agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt
     1141 taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggtgaa
     1201 agatccgcgt attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat
     1261 cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg
     1321 tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa
     1381 caacaataac aataacaaca acagcagcgg cagactggaa gttctgttcc aggggccgat
     1441 ggctgagatc ggaaccggtt tcccattcga cccgcattac gtcgaggttc tgggagaacg
     1501 catgcactac gtggatgtcg gaccacgtga cggtaccccg gtgttgttcc tgcatggtaa
     1561 ccctactagt tcgtacgtct ggaggaacat cattcctcat gttgccccca ctcacagatg
     1621 tatcgcacct gacctcattg gcatgggaaa gagtgataaa cccgacttgg gttacttctt
     1681 cgatgaccac gtgcgtttca tggatgcatt catcgaggcg ctcggcttgg aggaagtcgt
     1741 tctggtcatt catgactggg gatctgcact gggattccac tgggcaaaga ggaacccaga
     1801 gagagtgaaa ggcatcgctt tcatggaatt catcaggcca attccgactt gggacgagtg
     1861 gcctgaattc gcgagagaaa cattccaggc tttccgcaca acggatgtgg gtcgtaagct
     1921 gatcattgac cagaacgtct tcatcgaggg tacactgcca atgggcgtgg tccgcccgct
     1981 gactgaggtg gaaatggatc attaccgtga gccattcctg aacccggtcg acagggaacc
     2041 cctctggaga ttccctaacg agttgcccat cgctggcgaa ccagccaaca ttgttgcact
     2101 cgtggaggaa tacatggact ggttgcacca aagccctgtt cccaagttgc tgttctgggg
     2161 aaccccaggt gtgctgatcc ctcctgccga agcagcgagg ttggcaaagt ctctgcccaa
     2221 ctgtaaagcc gtggacatcg gtccaggcct gaacctcttg caggaggata accctgacct
     2281 catcggttca gaaattgcta gatggctgtc aactctcgag atcagtggag gatccgctgg
     2341 ctccgctgct ggttctggcg cggccgcaag cgctatggcc acaacctgtt acattgcaca
     2401 agataaaaat atatcatcac gaacagtaaa actgtctgct tacataaaca gtaatacaag
     2461 gggtgttatg cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt
     2521 ggatgtacag agtgatatta ttgacacgcc cgggcgacgg atggtgatcc ccctggccag
     2581 tgcccgtctg ctgtcagata aagtctcccg tgaactttac ccggtggtgc atatcgggga
     2641 tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg ccggtctccg ttatcgggga
     2701 agaagtggct gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt
     2761 ctggggaata taaatgtcag gctcccttat acacagccag tggcgcgccg taatagaagc
     2821 ttgtcgagaa gtactagagg atcataatca gccataccac atttgtagag gttttacttg
     2881 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     2941 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     3001 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     3061 tatcttatca tgtctggatc tgatcactgc ttgagcctag gagatccgaa ccagataagt
     3121 gaaagcctgc atccgcagat cttaattaat aaaacacgat acattgttat tagtacattt
     3181 attaagcgct agattctgtg cgttgttgat ttacagacaa ttgttgtacg tattttaata
     3241 attcattaaa tttataatct ttagggtggt atgttagagc gaaaatcaaa tgattttcag
     3301 cgtctttata tctgaattta aatattaaat cctcaataga tttgtaaaat aggtttcgat
     3361 tagtttcaaa caagggttgt ttttccgaac cgatggctgg actatctaat ggattttcgc
     3421 tcaacgccac aaaacttgcc aaatcttgta gcagcaatct agctttgtcg atattcgttt
     3481 gtgttttgtt ttgtaataaa ggttcgacgt cgttcaaaat attatgcgct tttgtatttc
     3541 tttcatcact gtcgttagtg tacaattgac tcgacgtaaa cacgttaaat aacgcttgga
     3601 catatttaac atcgggcgtg ttagctttat taggccgatt atcgtcgtcg tcccaaccct
     3661 cgtcgttaga agttgcttcc gaagacgatt ttgccatagc cacacgacgc ctattaattg
     3721 tgtcggctaa cacgtccgcg atcaaatttg tagttgagct ttttgggcct catgggcctt
     3781 ccgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aacatggtca
     3841 tagctgtttc cttgcgtatt gggcgctctc cgcttcctcg ctcactgact cgctgcgctc
     3901 ggtcgttcgg gtaaagcctg gggtgcctaa tgagcaaaag gccagcaaaa ggccaggaac
     3961 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4021 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4081 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4141 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4201 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4261 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4321 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4381 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     4441 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4501 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4561 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     4621 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     4681 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     4741 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     4801 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     4861 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     4921 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     4981 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5041 cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5101 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5161 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5221 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5281 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5341 agttgctctt gcccggcgtc aacacgggat aataccgcgc cacatagcag aactttaaaa
     5401 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     5461 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     5521 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     5581 gcgacacgga aatgttgaat actcattact cttccttttt caatattatt gaagcattta
     5641 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     5701 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     5761 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     5821 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgg ccgctacagg
     5881 gcgctcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgtttcg gtgcgggcct
     5941 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     6001 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc gacgtaatac
     6061 gactcactat agggcgaatt ggcggaaggc cgtcaaggcc gtgacgttaa aactattaag
     6121 ccatccaatc gaccgttagt cgaatcagga ccgctggtgc gagaagccgc gaagtatggc
     6181 gaatgcatcg tataacgtgt ggagtccgct cattagagcg tcatgtttag acaagaaagc
     6241 tacatattta attgatcccg atgattttat tgataaattg accctaactc catacacggt
     6301 attctacaat ggcggggttt tggtcaaaat ttccggactg cgattgtaca tgctgttaac
     6361 ggctccgccc actattaatg aaattaaaaa ttccaatttt aaaaaacgca gcaagagaaa
     6421 catttgtatg aaagaatgcg tagaaggaaa gaaaaatgtc gtggacatgc tgaacaacaa
     6481 gattaatatg cctccgtgta taaaaaaaat attgaacgat ttgaaagaaa acaatgtacc
     6541 gcgcggcggt atgtacagga agaggtttat actaaactgt tacattgcaa acgtggtttc
     6601 gtgtgccaag tgtgaaaacc gatgtttaat caaggctctg acgcatttct acaaccacga
     6661 ctccaagtgt gtgggtgaag tcatgcatct tttaatcaaa tcccaagatg tgtataaacc
     6721 accaaactgc caaaaaatga aaactgtgga caagctctgt ccgtttgctg gcaactgcaa
     6781 gggtctcaat cctatttgta attattgaat aataaaacaa ttataaatgc taaatttgtt
     6841 ttttattaac gatacaaacc aaacgcaaca agaacatttg tagtattatc tataattgaa
     6901 aacgcgtagt tataatcgct gaggtaatat ttaaaatcat tttcaaatga ttcacagtta
     6961 atttgcgaca atataatttt attttcacat aaactagacg ccttgtcgtc ttcttcttcg
     7021 tattccttct ctttttcatt tttctcctca taaaaattaa catagttatt atcgtatcca
     7081 tatatgtatc tatcgtatag agtaaatttt ttgttgtcat aaatatatat gtctttttta
     7141 atggggtgta tagtaccgct gcgcatagtt tttctgtaat ttacaacagt gctattttct
     7201 ggtagttctt cggagtgtgt tgctttaatt attaaattta tataatcaat gaatttggga
     7261 tcgtcggttt tgtacaatat gttgccggca tagtacgcag cttcttctag ttcaattaca
     7321 ccatttttta gcagcaccgg attaacataa ctttccaaaa tgttgtacga accgttaaac
     7381 aaaaacagtt cacctccctt ttctatacta ttgtctgcga gcagttgttt gttgttaaaa
     7441 ataacagcca ttgtaatgag acgcacaaac taatatcaca
//
