LOCUS       sequence-232332-        6156 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 mCitrine, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6156)
  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 6156)
  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: pOCC230
FEATURES             Location/Qualifiers
     source          1..6156
                     /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             222..527
                     /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     406..425
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             583..603
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(604..606,607..609,610..1323)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY*"
     primer_bind     complement(649..670)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(910..929)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1257..1278
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1453..1587
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1490..1509)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1544..1563
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1660..2032
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2384..2401)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2494..3082)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2574..2593)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3253..4044,4045..4113))
                     /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     3876..3895
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4114..4218)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4519..4541
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4533..4550
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4534..4550
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4561..4580
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4561..4579
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5190..5314
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5352..5957)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     promoter        6065..6156
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     6081..6098
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 attccggatt attcataccg tcccaccatc gggcgcggat cgccaccatg ggcagcagcc
       61 atcaccatca ccatcacagc agcggcagac tggaagttct gttccagggg ccggcggccg
      121 caagcgctat ggccacaacc tgttacattg cacaagataa aaatatatca tcacgaacag
      181 taaaactgtc tgcttacata aacagtaata caaggggtgt tatgcagttt aaggtttaca
      241 cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca
      301 cgcccgggcg acggatggtg atccccctgg ccagtgcccg tctgctgtca gataaagtct
      361 cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg
      421 atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg
      481 aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggctccc
      541 ttatacacag ccagtggcgc gccgggatcc agcagcggca gagaaaactt gtatttccag
      601 ggcatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      661 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      721 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      781 accctcgtga ccaccttcgg ctacggcctg atgtgcttcg cccgctaccc cgaccacatg
      841 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      901 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      961 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1021 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
     1081 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
     1141 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1201 cactacctga gctaccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     1261 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtactaa
     1321 tagaagcttg tcgagaagta ctagaggatc ataatcagcc ataccacatt tgtagaggtt
     1381 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     1441 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     1501 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     1561 atcaatgtat cttatcatgt ctggatctga tcactgcttg agcctaggag atccgaacca
     1621 gataagtgaa agcctgcatc cgcagatctt aattaataaa acacgataca ttgttattag
     1681 tacatttatt aagcgctaga ttctgtgcgt tgttgattta cagacaattg ttgtacgtat
     1741 tttaataatt cattaaattt ataatcttta gggtggtatg ttagagcgaa aatcaaatga
     1801 ttttcagcgt ctttatatct gaatttaaat attaaatcct caatagattt gtaaaatagg
     1861 tttcgattag tttcaaacaa gggttgtttt tccgaaccga tggctggact atctaatgga
     1921 ttttcgctca acgccacaaa acttgccaaa tcttgtagca gcaatctagc tttgtcgata
     1981 ttcgtttgtg ttttgttttg taataaaggt tcgacgtcgt tcaaaatatt atgcgctttt
     2041 gtatttcttt catcactgtc gttagtgtac aattgactcg acgtaaacac gttaaataac
     2101 gcttggacat atttaacatc gggcgtgtta gctttattag gccgattatc gtcgtcgtcc
     2161 caaccctcgt cgttagaagt tgcttccgaa gacgattttg ccatagccac acgacgccta
     2221 ttaattgtgt cggctaacac gtccgcgatc aaatttgtag ttgagctttt tgggcctcat
     2281 gggccttccg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaac
     2341 atggtcatag ctgtttcctt gcgtattggg cgctctccgc ttcctcgctc actgactcgc
     2401 tgcgctcggt cgttcgggta aagcctgggg tgcctaatga gcaaaaggcc agcaaaaggc
     2461 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2521 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2581 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2641 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     2701 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2761 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2821 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2881 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     2941 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     3001 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     3061 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     3121 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     3181 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     3241 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     3301 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     3361 accatctggc cccagtgctg caatgatacc gcgagaacca cgctcaccgg ctccagattt
     3421 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3481 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3541 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3601 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3661 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3721 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3781 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3841 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3901 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3961 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     4021 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     4081 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     4141 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     4201 acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat
     4261 attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc
     4321 gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtggccgc
     4381 tacagggcgc tcccattcgc cattcaggct gcgcaactgt tgggaagggc gtttcggtgc
     4441 gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt
     4501 gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt gagcgcgacg
     4561 taatacgact cactataggg cgaattggcg gaaggccgtc aaggccgtga cgttaaaact
     4621 attaagccat ccaatcgacc gttagtcgaa tcaggaccgc tggtgcgaga agccgcgaag
     4681 tatggcgaat gcatcgtata acgtgtggag tccgctcatt agagcgtcat gtttagacaa
     4741 gaaagctaca tatttaattg atcccgatga ttttattgat aaattgaccc taactccata
     4801 cacggtattc tacaatggcg gggttttggt caaaatttcc ggactgcgat tgtacatgct
     4861 gttaacggct ccgcccacta ttaatgaaat taaaaattcc aattttaaaa aacgcagcaa
     4921 gagaaacatt tgtatgaaag aatgcgtaga aggaaagaaa aatgtcgtgg acatgctgaa
     4981 caacaagatt aatatgcctc cgtgtataaa aaaaatattg aacgatttga aagaaaacaa
     5041 tgtaccgcgc ggcggtatgt acaggaagag gtttatacta aactgttaca ttgcaaacgt
     5101 ggtttcgtgt gccaagtgtg aaaaccgatg tttaatcaag gctctgacgc atttctacaa
     5161 ccacgactcc aagtgtgtgg gtgaagtcat gcatctttta atcaaatccc aagatgtgta
     5221 taaaccacca aactgccaaa aaatgaaaac tgtggacaag ctctgtccgt ttgctggcaa
     5281 ctgcaagggt ctcaatccta tttgtaatta ttgaataata aaacaattat aaatgctaaa
     5341 tttgtttttt attaacgata caaaccaaac gcaacaagaa catttgtagt attatctata
     5401 attgaaaacg cgtagttata atcgctgagg taatatttaa aatcattttc aaatgattca
     5461 cagttaattt gcgacaatat aattttattt tcacataaac tagacgcctt gtcgtcttct
     5521 tcttcgtatt ccttctcttt ttcatttttc tcctcataaa aattaacata gttattatcg
     5581 tatccatata tgtatctatc gtatagagta aattttttgt tgtcataaat atatatgtct
     5641 tttttaatgg ggtgtatagt accgctgcgc atagtttttc tgtaatttac aacagtgcta
     5701 ttttctggta gttcttcgga gtgtgttgct ttaattatta aatttatata atcaatgaat
     5761 ttgggatcgt cggttttgta caatatgttg ccggcatagt acgcagcttc ttctagttca
     5821 attacaccat tttttagcag caccggatta acataacttt ccaaaatgtt gtacgaaccg
     5881 ttaaacaaaa acagttcacc tcccttttct atactattgt ctgcgagcag ttgtttgttg
     5941 ttaaaaataa cagccattgt aatgagacgc acaaactaat atcacaagct gccaccattt
     6001 tgttcgccca ggactctagc tatagttcta gtggttggct acgtatactc cggaatatta
     6061 atagatcatg gagataatta aaatgataac catctcgcaa ataaataagt attttactgt
     6121 tttcgtaaca gttttgtaat aaaaaaacct ataaat
//
