LOCUS       sequence-232236-        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,
            and C-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: pOCC139
FEATURES             Location/Qualifiers
     source          1..6150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..24
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             133..438
                     /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     317..336
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             join(506..508,509..511,512..1225)
                     /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(551..572)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(812..831)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1159..1180
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1358..1492
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1395..1414)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1449..1468
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1565..1937
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2289..2306)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2399..2987)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2479..2498)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3158..3949,3950..4018))
                     /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     3781..3800
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4019..4123)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4424..4446
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4438..4455
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4439..4455
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4466..4485
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4466..4484
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5095..5219
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5257..5862)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     promoter        5970..6061
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     5986..6003
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     regulatory      6103..6112
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             6121..6138
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 ctggaagttc tgttccaggg gccggcggcc gcaagcgcta tggccacaac ctgttacatt
       61 gcacaagata aaaatatatc atcacgaaca gtaaaactgt ctgcttacat aaacagtaat
      121 acaaggggtg ttatgcagtt taaggtttac acctataaaa gagagagccg ttatcgtctg
      181 tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt gatccccctg
      241 gccagtgccc gtctgctgtc agataaagtc tcccgtgaac tttacccggt ggtgcatatc
      301 ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt ctccgttatc
      361 ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg
      421 atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtggcg cgccgggatc
      481 cgctggctcc gctgctggtt ctggcatggt gagcaagggc gaggagctgt tcaccggggt
      541 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
      601 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
      661 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
      721 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
      781 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
      841 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
      901 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
      961 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     1021 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     1081 ccccgtgctg ctgcccgaca accactacct gagcacccag tccaaactga gcaaagaccc
     1141 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     1201 cggcatggac gagctgtaca agtaatagaa gcttgtcgag aagtactaga ggatcataat
     1261 cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac acctccccct
     1321 gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg cagcttataa
     1381 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     1441 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga tctgatcact
     1501 gcttgagcct aggagatccg aaccagataa gtgaaagcct gcatccgcag atcttaatta
     1561 ataaaacacg atacattgtt attagtacat ttattaagcg ctagattctg tgcgttgttg
     1621 atttacagac aattgttgta cgtattttaa taattcatta aatttataat ctttagggtg
     1681 gtatgttaga gcgaaaatca aatgattttc agcgtcttta tatctgaatt taaatattaa
     1741 atcctcaata gatttgtaaa ataggtttcg attagtttca aacaagggtt gtttttccga
     1801 accgatggct ggactatcta atggattttc gctcaacgcc acaaaacttg ccaaatcttg
     1861 tagcagcaat ctagctttgt cgatattcgt ttgtgttttg ttttgtaata aaggttcgac
     1921 gtcgttcaaa atattatgcg cttttgtatt tctttcatca ctgtcgttag tgtacaattg
     1981 actcgacgta aacacgttaa ataacgcttg gacatattta acatcgggcg tgttagcttt
     2041 attaggccga ttatcgtcgt cgtcccaacc ctcgtcgtta gaagttgctt ccgaagacga
     2101 ttttgccata gccacacgac gcctattaat tgtgtcggct aacacgtccg cgatcaaatt
     2161 tgtagttgag ctttttgggc ctcatgggcc ttccgctcac tgcccgcttt ccagtcggga
     2221 aacctgtcgt gccagctgca ttaacatggt catagctgtt tccttgcgta ttgggcgctc
     2281 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc gggtaaagcc tggggtgcct
     2341 aatgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     2401 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     2461 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     2521 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2581 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2641 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2701 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2761 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2821 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2881 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2941 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     3001 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     3061 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     3121 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     3181 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     3241 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     3301 aaccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     3361 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     3421 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     3481 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     3541 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     3601 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     3661 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     3721 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     3781 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     3841 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     3901 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     3961 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     4021 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     4081 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     4141 tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     4201 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     4261 gaccgagata gggttgagtg gccgctacag ggcgctccca ttcgccattc aggctgcgca
     4321 actgttggga agggcgtttc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg
     4381 ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt
     4441 aaaacgacgg ccagtgagcg cgacgtaata cgactcacta tagggcgaat tggcggaagg
     4501 ccgtcaaggc cgtgacgtta aaactattaa gccatccaat cgaccgttag tcgaatcagg
     4561 accgctggtg cgagaagccg cgaagtatgg cgaatgcatc gtataacgtg tggagtccgc
     4621 tcattagagc gtcatgttta gacaagaaag ctacatattt aattgatccc gatgatttta
     4681 ttgataaatt gaccctaact ccatacacgg tattctacaa tggcggggtt ttggtcaaaa
     4741 tttccggact gcgattgtac atgctgttaa cggctccgcc cactattaat gaaattaaaa
     4801 attccaattt taaaaaacgc agcaagagaa acatttgtat gaaagaatgc gtagaaggaa
     4861 agaaaaatgt cgtggacatg ctgaacaaca agattaatat gcctccgtgt ataaaaaaaa
     4921 tattgaacga tttgaaagaa aacaatgtac cgcgcggcgg tatgtacagg aagaggttta
     4981 tactaaactg ttacattgca aacgtggttt cgtgtgccaa gtgtgaaaac cgatgtttaa
     5041 tcaaggctct gacgcatttc tacaaccacg actccaagtg tgtgggtgaa gtcatgcatc
     5101 ttttaatcaa atcccaagat gtgtataaac caccaaactg ccaaaaaatg aaaactgtgg
     5161 acaagctctg tccgtttgct ggcaactgca agggtctcaa tcctatttgt aattattgaa
     5221 taataaaaca attataaatg ctaaatttgt tttttattaa cgatacaaac caaacgcaac
     5281 aagaacattt gtagtattat ctataattga aaacgcgtag ttataatcgc tgaggtaata
     5341 tttaaaatca ttttcaaatg attcacagtt aatttgcgac aatataattt tattttcaca
     5401 taaactagac gccttgtcgt cttcttcttc gtattccttc tctttttcat ttttctcctc
     5461 ataaaaatta acatagttat tatcgtatcc atatatgtat ctatcgtata gagtaaattt
     5521 tttgttgtca taaatatata tgtctttttt aatggggtgt atagtaccgc tgcgcatagt
     5581 ttttctgtaa tttacaacag tgctattttc tggtagttct tcggagtgtg ttgctttaat
     5641 tattaaattt atataatcaa tgaatttggg atcgtcggtt ttgtacaata tgttgccggc
     5701 atagtacgca gcttcttcta gttcaattac accatttttt agcagcaccg gattaacata
     5761 actttccaaa atgttgtacg aaccgttaaa caaaaacagt tcacctccct tttctatact
     5821 attgtctgcg agcagttgtt tgttgttaaa aataacagcc attgtaatga gacgcacaaa
     5881 ctaatatcac aagctgccac cattttgttc gcccaggact ctagctatag ttctagtggt
     5941 tggctacgta tactccggaa tattaataga tcatggagat aattaaaatg ataaccatct
     6001 cgcaaataaa taagtatttt actgttttcg taacagtttt gtaataaaaa aacctataaa
     6061 tattccggat tattcatacc gtcccaccat cgggcgcgga tcgccaccat gggcagcagc
     6121 catcaccatc accatcacag cagcggcaga
//
