LOCUS       sequence-232317-        6150 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 
            protease, and N-terminal monomeric GFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6150)
  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 6150)
  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: pOCC65
FEATURES             Location/Qualifiers
     source          1..6150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      42..51
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             60..77
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             90..113
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(114..116,117..119,120..830)
                     /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(159..180)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(420..439)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     767..788
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             969..1274
                     /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     1153..1172
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    1447..1581
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1484..1503)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1538..1557
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1654..2026
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2378..2395)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2488..3076)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2568..2587)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3247..4038,4039..4107))
                     /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     3870..3889
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4108..4212)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4513..4535
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4527..4544
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4528..4544
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4555..4574
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4555..4573
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5184..5308
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5346..5951)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     promoter        6059..6150
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     6075..6092
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 attccggatt attcataccg tcccaccatc gggcgcggat cgccaccatg ggcagcagcc
       61 atcaccatca ccatcacagc agcggcagac tggaagttct gttccagggg ccgatggtga
      121 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg
      181 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc
      241 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga
      301 ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg
      361 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg
      421 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc
      481 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg
      541 agtacaacta caacagccac aacgtctata tcatggccga caagcagaag aacggcatca
      601 aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact
      661 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga
      721 gcacccagtc caaactgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg
      781 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag ggatccgctg
      841 gctccgctgc tggttctggc gcggccgcaa gcgctatggc cacaacctgt tacattgcac
      901 aagataaaaa tatatcatca cgaacagtaa aactgtctgc ttacataaac agtaatacaa
      961 ggggtgttat gcagtttaag gtttacacct ataaaagaga gagccgttat cgtctgtttg
     1021 tggatgtaca gagtgatatt attgacacgc ccgggcgacg gatggtgatc cccctggcca
     1081 gtgcccgtct gctgtcagat aaagtctccc gtgaacttta cccggtggtg catatcgggg
     1141 atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggtctcc gttatcgggg
     1201 aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt aacctgatgt
     1261 tctggggaat ataaatgtca ggctccctta tacacagcca gtggcgcgcc gtaatagaag
     1321 cttgtcgaga agtactagag gatcataatc agccatacca catttgtaga ggttttactt
     1381 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     1441 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     1501 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     1561 gtatcttatc atgtctggat ctgatcactg cttgagccta ggagatccga accagataag
     1621 tgaaagcctg catccgcaga tcttaattaa taaaacacga tacattgtta ttagtacatt
     1681 tattaagcgc tagattctgt gcgttgttga tttacagaca attgttgtac gtattttaat
     1741 aattcattaa atttataatc tttagggtgg tatgttagag cgaaaatcaa atgattttca
     1801 gcgtctttat atctgaattt aaatattaaa tcctcaatag atttgtaaaa taggtttcga
     1861 ttagtttcaa acaagggttg tttttccgaa ccgatggctg gactatctaa tggattttcg
     1921 ctcaacgcca caaaacttgc caaatcttgt agcagcaatc tagctttgtc gatattcgtt
     1981 tgtgttttgt tttgtaataa aggttcgacg tcgttcaaaa tattatgcgc ttttgtattt
     2041 ctttcatcac tgtcgttagt gtacaattga ctcgacgtaa acacgttaaa taacgcttgg
     2101 acatatttaa catcgggcgt gttagcttta ttaggccgat tatcgtcgtc gtcccaaccc
     2161 tcgtcgttag aagttgcttc cgaagacgat tttgccatag ccacacgacg cctattaatt
     2221 gtgtcggcta acacgtccgc gatcaaattt gtagttgagc tttttgggcc tcatgggcct
     2281 tccgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taacatggtc
     2341 atagctgttt ccttgcgtat tgggcgctct ccgcttcctc gctcactgac tcgctgcgct
     2401 cggtcgttcg ggtaaagcct ggggtgccta atgagcaaaa ggccagcaaa aggccaggaa
     2461 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2521 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2581 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     2641 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     2701 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     2761 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     2821 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     2881 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     2941 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3001 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3061 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3121 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     3181 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     3241 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     3301 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     3361 tggccccagt gctgcaatga taccgcgaga accacgctca ccggctccag atttatcagc
     3421 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     3481 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     3541 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     3601 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     3661 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     3721 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     3781 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     3841 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     3901 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     3961 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     4021 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     4081 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     4141 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     4201 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     4261 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     4321 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgg ccgctacagg
     4381 gcgctcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgtttcg gtgcgggcct
     4441 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     4501 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgagcgc gacgtaatac
     4561 gactcactat agggcgaatt ggcggaaggc cgtcaaggcc gtgacgttaa aactattaag
     4621 ccatccaatc gaccgttagt cgaatcagga ccgctggtgc gagaagccgc gaagtatggc
     4681 gaatgcatcg tataacgtgt ggagtccgct cattagagcg tcatgtttag acaagaaagc
     4741 tacatattta attgatcccg atgattttat tgataaattg accctaactc catacacggt
     4801 attctacaat ggcggggttt tggtcaaaat ttccggactg cgattgtaca tgctgttaac
     4861 ggctccgccc actattaatg aaattaaaaa ttccaatttt aaaaaacgca gcaagagaaa
     4921 catttgtatg aaagaatgcg tagaaggaaa gaaaaatgtc gtggacatgc tgaacaacaa
     4981 gattaatatg cctccgtgta taaaaaaaat attgaacgat ttgaaagaaa acaatgtacc
     5041 gcgcggcggt atgtacagga agaggtttat actaaactgt tacattgcaa acgtggtttc
     5101 gtgtgccaag tgtgaaaacc gatgtttaat caaggctctg acgcatttct acaaccacga
     5161 ctccaagtgt gtgggtgaag tcatgcatct tttaatcaaa tcccaagatg tgtataaacc
     5221 accaaactgc caaaaaatga aaactgtgga caagctctgt ccgtttgctg gcaactgcaa
     5281 gggtctcaat cctatttgta attattgaat aataaaacaa ttataaatgc taaatttgtt
     5341 ttttattaac gatacaaacc aaacgcaaca agaacatttg tagtattatc tataattgaa
     5401 aacgcgtagt tataatcgct gaggtaatat ttaaaatcat tttcaaatga ttcacagtta
     5461 atttgcgaca atataatttt attttcacat aaactagacg ccttgtcgtc ttcttcttcg
     5521 tattccttct ctttttcatt tttctcctca taaaaattaa catagttatt atcgtatcca
     5581 tatatgtatc tatcgtatag agtaaatttt ttgttgtcat aaatatatat gtctttttta
     5641 atggggtgta tagtaccgct gcgcatagtt tttctgtaat ttacaacagt gctattttct
     5701 ggtagttctt cggagtgtgt tgctttaatt attaaattta tataatcaat gaatttggga
     5761 tcgtcggttt tgtacaatat gttgccggca tagtacgcag cttcttctag ttcaattaca
     5821 ccatttttta gcagcaccgg attaacataa ctttccaaaa tgttgtacga accgttaaac
     5881 aaaaacagtt cacctccctt ttctatacta ttgtctgcga gcagttgttt gttgttaaaa
     5941 ataacagcca ttgtaatgag acgcacaaac taatatcaca agctgccacc attttgttcg
     6001 cccaggactc tagctatagt tctagtggtt ggctacgtat actccggaat attaatagat
     6061 catggagata attaaaatga taaccatctc gcaaataaat aagtatttta ctgttttcgt
     6121 aacagttttg taataaaaaa acctataaat
//
