LOCUS       sequence-224700-        6183 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with honey bee melitine secretion signal and
            C-terminal eGFP-HIS6.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6183)
  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 6183)
  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: pOCC92
FEATURES             Location/Qualifiers
     source          1..6183
                     /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"
     sig_peptide     224..283
                     /codon_start=1
                     /product="signal sequence from honeybee melittin"
                     /label=melittin signal sequence
                     /translation="KFLVNVALVFMVVYISYIYA"
     CDS             404..709
                     /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     588..607
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             join(771..773,774..776,777..1487)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(771..773,774..776,777..1487)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(816..837)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1077..1096)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1424..1445
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1497..1514
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1649..1783
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1686..1705)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1740..1759
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1856..2228
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2580..2597)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2690..3278)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2770..2789)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3449..4240,4241..4309))
                     /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     4072..4091
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4311..4415)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4716..4738
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4730..4747
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4731..4747
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4758..4777
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4758..4776
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5387..5511
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5549..6154)
                     /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 ggcaaattct tagtcaacgt
      241 tgcccttgtt tttatggtcg tatacatttc ttacatctat gcgggatcca attgggcggc
      301 cgcaagcgct atggccacaa cctgttacat tgcacaagat aaaaatatat catcacgaac
      361 agtaaaactg tctgcttaca taaacagtaa tacaaggggt gttatgcagt ttaaggttta
      421 cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga
      481 cacgcccggg cgacggatgg tgatccccct ggccagtgcc cgtctgctgt cagataaagt
      541 ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac
      601 cgatatggcc agtgtgccgg tctccgttat cggggaagaa gtggctgatc tcagccaccg
      661 cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggctc
      721 ccttatacac agccagtggc gcgccggctg gctccgctgc tggttctggc atggtgagca
      781 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
      841 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
      901 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
      961 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     1021 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     1081 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     1141 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     1201 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     1261 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     1321 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     1381 cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     1441 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagagc agcggccatc
     1501 accatcacca tcactaaaga agcttgtcga gaagtactag aggatcataa tcagccatac
     1561 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     1621 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     1681 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     1741 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atctgatcac tgcttgagcc
     1801 taggagatcc gaaccagata agtgaaagcc tgcatccgca gatcttaatt aataaaacac
     1861 gatacattgt tattagtaca tttattaagc gctagattct gtgcgttgtt gatttacaga
     1921 caattgttgt acgtatttta ataattcatt aaatttataa tctttagggt ggtatgttag
     1981 agcgaaaatc aaatgatttt cagcgtcttt atatctgaat ttaaatatta aatcctcaat
     2041 agatttgtaa aataggtttc gattagtttc aaacaagggt tgtttttccg aaccgatggc
     2101 tggactatct aatggatttt cgctcaacgc cacaaaactt gccaaatctt gtagcagcaa
     2161 tctagctttg tcgatattcg tttgtgtttt gttttgtaat aaaggttcga cgtcgttcaa
     2221 aatattatgc gcttttgtat ttctttcatc actgtcgtta gtgtacaatt gactcgacgt
     2281 aaacacgtta aataacgctt ggacatattt aacatcgggc gtgttagctt tattaggccg
     2341 attatcgtcg tcgtcccaac cctcgtcgtt agaagttgct tccgaagacg attttgccat
     2401 agccacacga cgcctattaa ttgtgtcggc taacacgtcc gcgatcaaat ttgtagttga
     2461 gctttttggg cctcatgggc cttccgctca ctgcccgctt tccagtcggg aaacctgtcg
     2521 tgccagctgc attaacatgg tcatagctgt ttccttgcgt attgggcgct ctccgcttcc
     2581 tcgctcactg actcgctgcg ctcggtcgtt cgggtaaagc ctggggtgcc taatgagcaa
     2641 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     2701 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     2761 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     2821 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     2881 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     2941 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     3001 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     3061 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     3121 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     3181 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     3241 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     3301 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     3361 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3421 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     3481 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     3541 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     3601 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     3661 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     3721 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctgcaggc atcgtggtgt
     3781 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     3841 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     3901 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     3961 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     4021 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaacacgg gataataccg
     4081 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     4141 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     4201 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     4261 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcatt actcttcctt
     4321 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4381 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4441 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     4501 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     4561 tagggttgag tggccgctac agggcgctcc cattcgccat tcaggctgcg caactgttgg
     4621 gaagggcgtt tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     4681 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     4741 ggccagtgag cgcgacgtaa tacgactcac tatagggcga attggcggaa ggccgtcaag
     4801 gccgtgacgt taaaactatt aagccatcca atcgaccgtt agtcgaatca ggaccgctgg
     4861 tgcgagaagc cgcgaagtat ggcgaatgca tcgtataacg tgtggagtcc gctcattaga
     4921 gcgtcatgtt tagacaagaa agctacatat ttaattgatc ccgatgattt tattgataaa
     4981 ttgaccctaa ctccatacac ggtattctac aatggcgggg ttttggtcaa aatttccgga
     5041 ctgcgattgt acatgctgtt aacggctccg cccactatta atgaaattaa aaattccaat
     5101 tttaaaaaac gcagcaagag aaacatttgt atgaaagaat gcgtagaagg aaagaaaaat
     5161 gtcgtggaca tgctgaacaa caagattaat atgcctccgt gtataaaaaa aatattgaac
     5221 gatttgaaag aaaacaatgt accgcgcggc ggtatgtaca ggaagaggtt tatactaaac
     5281 tgttacattg caaacgtggt ttcgtgtgcc aagtgtgaaa accgatgttt aatcaaggct
     5341 ctgacgcatt tctacaacca cgactccaag tgtgtgggtg aagtcatgca tcttttaatc
     5401 aaatcccaag atgtgtataa accaccaaac tgccaaaaaa tgaaaactgt ggacaagctc
     5461 tgtccgtttg ctggcaactg caagggtctc aatcctattt gtaattattg aataataaaa
     5521 caattataaa tgctaaattt gttttttatt aacgatacaa accaaacgca acaagaacat
     5581 ttgtagtatt atctataatt gaaaacgcgt agttataatc gctgaggtaa tatttaaaat
     5641 cattttcaaa tgattcacag ttaatttgcg acaatataat tttattttca cataaactag
     5701 acgccttgtc gtcttcttct tcgtattcct tctctttttc atttttctcc tcataaaaat
     5761 taacatagtt attatcgtat ccatatatgt atctatcgta tagagtaaat tttttgttgt
     5821 cataaatata tatgtctttt ttaatggggt gtatagtacc gctgcgcata gtttttctgt
     5881 aatttacaac agtgctattt tctggtagtt cttcggagtg tgttgcttta attattaaat
     5941 ttatataatc aatgaatttg ggatcgtcgg ttttgtacaa tatgttgccg gcatagtacg
     6001 cagcttcttc tagttcaatt acaccatttt ttagcagcac cggattaaca taactttcca
     6061 aaatgttgta cgaaccgtta aacaaaaaca gttcacctcc cttttctata ctattgtctg
     6121 cgagcagttg tttgttgtta aaaataacag ccattgtaat gagacgcaca aactaatatc
     6181 aca
//
