LOCUS       sequence-224664-        6160 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal HIS6 tag, cleavable with 3C,
            and C-terminal monomeric GFP, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6160)
  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 6160)
  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: pOCC69
FEATURES             Location/Qualifiers
     source          1..6160
                     /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..283
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             392..697
                     /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     576..595
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             753..773
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(774..776,777..779,780..1493)
                     /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(819..840)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1080..1099)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1427..1448
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1626..1760
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1663..1682)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1717..1736
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1833..2205
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2557..2574)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2667..3255)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2747..2766)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3426..4217,4218..4286))
                     /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     4049..4068
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4288..4392)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4693..4715
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4707..4724
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4708..4724
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4735..4754
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4735..4753
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5364..5488
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5526..6131)
                     /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 agcggcagac tggaagttct gttccagggg ccggcggccg caagcgctat
      301 ggccacaacc tgttacattg cacaagataa aaatatatca tcacgaacag taaaactgtc
      361 tgcttacata aacagtaata caaggggtgt tatgcagttt aaggtttaca cctataaaag
      421 agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca cgcccgggcg
      481 acggatggtg atccccctgg ccagtgcccg tctgctgtca gataaagtct cccgtgaact
      541 ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag
      601 tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat
      661 caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggctccc ttatacacag
      721 ccagtggcgc gccgggatcc agcagcggca gagaaaactt gtatttccag ggcatggtga
      781 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
      841 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
      901 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
      961 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
     1021 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
     1081 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
     1141 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
     1201 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
     1261 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
     1321 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
     1381 gcacccagtc caaactgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
     1441 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taatagaagc
     1501 ttgtcgagaa gtactagagg atcataatca gccataccac atttgtagag gttttacttg
     1561 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     1621 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1681 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     1741 tatcttatca tgtctggatc tgatcactgc ttgagcctag gagatccgaa ccagataagt
     1801 gaaagcctgc atccgcagat cttaattaat aaaacacgat acattgttat tagtacattt
     1861 attaagcgct agattctgtg cgttgttgat ttacagacaa ttgttgtacg tattttaata
     1921 attcattaaa tttataatct ttagggtggt atgttagagc gaaaatcaaa tgattttcag
     1981 cgtctttata tctgaattta aatattaaat cctcaataga tttgtaaaat aggtttcgat
     2041 tagtttcaaa caagggttgt ttttccgaac cgatggctgg actatctaat ggattttcgc
     2101 tcaacgccac aaaacttgcc aaatcttgta gcagcaatct agctttgtcg atattcgttt
     2161 gtgttttgtt ttgtaataaa ggttcgacgt cgttcaaaat attatgcgct tttgtatttc
     2221 tttcatcact gtcgttagtg tacaattgac tcgacgtaaa cacgttaaat aacgcttgga
     2281 catatttaac atcgggcgtg ttagctttat taggccgatt atcgtcgtcg tcccaaccct
     2341 cgtcgttaga agttgcttcc gaagacgatt ttgccatagc cacacgacgc ctattaattg
     2401 tgtcggctaa cacgtccgcg atcaaatttg tagttgagct ttttgggcct catgggcctt
     2461 ccgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aacatggtca
     2521 tagctgtttc cttgcgtatt gggcgctctc cgcttcctcg ctcactgact cgctgcgctc
     2581 ggtcgttcgg gtaaagcctg gggtgcctaa tgagcaaaag gccagcaaaa ggccaggaac
     2641 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     2701 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     2761 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     2821 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     2881 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     2941 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     3001 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     3061 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     3121 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     3181 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     3241 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     3301 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     3361 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     3421 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     3481 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     3541 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     3601 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     3661 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     3721 cgcaacgttg ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     3781 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     3841 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     3901 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     3961 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     4021 agttgctctt gcccggcgtc aacacgggat aataccgcgc cacatagcag aactttaaaa
     4081 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     4141 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     4201 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     4261 gcgacacgga aatgttgaat actcattact cttccttttt caatattatt gaagcattta
     4321 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     4381 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     4441 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     4501 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgg ccgctacagg
     4561 gcgctcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgtttcg gtgcgggcct
     4621 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     4681 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc gacgtaatac
     4741 gactcactat agggcgaatt ggcggaaggc cgtcaaggcc gtgacgttaa aactattaag
     4801 ccatccaatc gaccgttagt cgaatcagga ccgctggtgc gagaagccgc gaagtatggc
     4861 gaatgcatcg tataacgtgt ggagtccgct cattagagcg tcatgtttag acaagaaagc
     4921 tacatattta attgatcccg atgattttat tgataaattg accctaactc catacacggt
     4981 attctacaat ggcggggttt tggtcaaaat ttccggactg cgattgtaca tgctgttaac
     5041 ggctccgccc actattaatg aaattaaaaa ttccaatttt aaaaaacgca gcaagagaaa
     5101 catttgtatg aaagaatgcg tagaaggaaa gaaaaatgtc gtggacatgc tgaacaacaa
     5161 gattaatatg cctccgtgta taaaaaaaat attgaacgat ttgaaagaaa acaatgtacc
     5221 gcgcggcggt atgtacagga agaggtttat actaaactgt tacattgcaa acgtggtttc
     5281 gtgtgccaag tgtgaaaacc gatgtttaat caaggctctg acgcatttct acaaccacga
     5341 ctccaagtgt gtgggtgaag tcatgcatct tttaatcaaa tcccaagatg tgtataaacc
     5401 accaaactgc caaaaaatga aaactgtgga caagctctgt ccgtttgctg gcaactgcaa
     5461 gggtctcaat cctatttgta attattgaat aataaaacaa ttataaatgc taaatttgtt
     5521 ttttattaac gatacaaacc aaacgcaaca agaacatttg tagtattatc tataattgaa
     5581 aacgcgtagt tataatcgct gaggtaatat ttaaaatcat tttcaaatga ttcacagtta
     5641 atttgcgaca atataatttt attttcacat aaactagacg ccttgtcgtc ttcttcttcg
     5701 tattccttct ctttttcatt tttctcctca taaaaattaa catagttatt atcgtatcca
     5761 tatatgtatc tatcgtatag agtaaatttt ttgttgtcat aaatatatat gtctttttta
     5821 atggggtgta tagtaccgct gcgcatagtt tttctgtaat ttacaacagt gctattttct
     5881 ggtagttctt cggagtgtgt tgctttaatt attaaattta tataatcaat gaatttggga
     5941 tcgtcggttt tgtacaatat gttgccggca tagtacgcag cttcttctag ttcaattaca
     6001 ccatttttta gcagcaccgg attaacataa ctttccaaaa tgttgtacga accgttaaac
     6061 aaaaacagtt cacctccctt ttctatacta ttgtctgcga gcagttgttt gttgttaaaa
     6121 ataacagcca ttgtaatgag acgcacaaac taatatcaca
//
