LOCUS       sequence-232323-        6159 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 N-terminal monomeric GFP, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6159)
  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 6159)
  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: pOCC72
FEATURES             Location/Qualifiers
     source          1..6159
                     /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             849..869
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             978..1283
                     /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     1162..1181
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     polyA_signal    1456..1590
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1493..1512)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1547..1566
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           1663..2035
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(2387..2404)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2497..3085)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2577..2596)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3256..4047,4048..4116))
                     /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     3879..3898
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4117..4221)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4522..4544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4536..4553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4537..4553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4564..4583
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4564..4582
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5193..5317
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(5355..5960)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     promoter        6068..6159
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     6084..6101
                     /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 ggatccagca
      841 gcggcagaga aaacttgtat ttccagggcg cggccgcaag cgctatggcc acaacctgtt
      901 acattgcaca agataaaaat atatcatcac gaacagtaaa actgtctgct tacataaaca
      961 gtaatacaag gggtgttatg cagtttaagg tttacaccta taaaagagag agccgttatc
     1021 gtctgtttgt ggatgtacag agtgatatta ttgacacgcc cgggcgacgg atggtgatcc
     1081 ccctggccag tgcccgtctg ctgtcagata aagtctcccg tgaactttac ccggtggtgc
     1141 atatcgggga tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg ccggtctccg
     1201 ttatcgggga agaagtggct gatctcagcc accgcgaaaa tgacatcaaa aacgccatta
     1261 acctgatgtt ctggggaata taaatgtcag gctcccttat acacagccag tggcgcgccg
     1321 taatagaagc ttgtcgagaa gtactagagg atcataatca gccataccac atttgtagag
     1381 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     1441 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     1501 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     1561 ctcatcaatg tatcttatca tgtctggatc tgatcactgc ttgagcctag gagatccgaa
     1621 ccagataagt gaaagcctgc atccgcagat cttaattaat aaaacacgat acattgttat
     1681 tagtacattt attaagcgct agattctgtg cgttgttgat ttacagacaa ttgttgtacg
     1741 tattttaata attcattaaa tttataatct ttagggtggt atgttagagc gaaaatcaaa
     1801 tgattttcag cgtctttata tctgaattta aatattaaat cctcaataga tttgtaaaat
     1861 aggtttcgat tagtttcaaa caagggttgt ttttccgaac cgatggctgg actatctaat
     1921 ggattttcgc tcaacgccac aaaacttgcc aaatcttgta gcagcaatct agctttgtcg
     1981 atattcgttt gtgttttgtt ttgtaataaa ggttcgacgt cgttcaaaat attatgcgct
     2041 tttgtatttc tttcatcact gtcgttagtg tacaattgac tcgacgtaaa cacgttaaat
     2101 aacgcttgga catatttaac atcgggcgtg ttagctttat taggccgatt atcgtcgtcg
     2161 tcccaaccct cgtcgttaga agttgcttcc gaagacgatt ttgccatagc cacacgacgc
     2221 ctattaattg tgtcggctaa cacgtccgcg atcaaatttg tagttgagct ttttgggcct
     2281 catgggcctt ccgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     2341 aacatggtca tagctgtttc cttgcgtatt gggcgctctc cgcttcctcg ctcactgact
     2401 cgctgcgctc ggtcgttcgg gtaaagcctg gggtgcctaa tgagcaaaag gccagcaaaa
     2461 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     2521 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     2581 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     2641 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     2701 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     2761 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     2821 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     2881 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     2941 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3001 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3061 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3121 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3181 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3241 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     3301 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     3361 cttaccatct ggccccagtg ctgcaatgat accgcgagaa ccacgctcac cggctccaga
     3421 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     3481 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     3541 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     3601 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     3661 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     3721 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     3781 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     3841 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     3901 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     3961 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     4021 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     4081 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     4141 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     4201 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     4261 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     4321 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtggc
     4381 cgctacaggg cgctcccatt cgccattcag gctgcgcaac tgttgggaag ggcgtttcgg
     4441 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa
     4501 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgagcgcg
     4561 acgtaatacg actcactata gggcgaattg gcggaaggcc gtcaaggccg tgacgttaaa
     4621 actattaagc catccaatcg accgttagtc gaatcaggac cgctggtgcg agaagccgcg
     4681 aagtatggcg aatgcatcgt ataacgtgtg gagtccgctc attagagcgt catgtttaga
     4741 caagaaagct acatatttaa ttgatcccga tgattttatt gataaattga ccctaactcc
     4801 atacacggta ttctacaatg gcggggtttt ggtcaaaatt tccggactgc gattgtacat
     4861 gctgttaacg gctccgccca ctattaatga aattaaaaat tccaatttta aaaaacgcag
     4921 caagagaaac atttgtatga aagaatgcgt agaaggaaag aaaaatgtcg tggacatgct
     4981 gaacaacaag attaatatgc ctccgtgtat aaaaaaaata ttgaacgatt tgaaagaaaa
     5041 caatgtaccg cgcggcggta tgtacaggaa gaggtttata ctaaactgtt acattgcaaa
     5101 cgtggtttcg tgtgccaagt gtgaaaaccg atgtttaatc aaggctctga cgcatttcta
     5161 caaccacgac tccaagtgtg tgggtgaagt catgcatctt ttaatcaaat cccaagatgt
     5221 gtataaacca ccaaactgcc aaaaaatgaa aactgtggac aagctctgtc cgtttgctgg
     5281 caactgcaag ggtctcaatc ctatttgtaa ttattgaata ataaaacaat tataaatgct
     5341 aaatttgttt tttattaacg atacaaacca aacgcaacaa gaacatttgt agtattatct
     5401 ataattgaaa acgcgtagtt ataatcgctg aggtaatatt taaaatcatt ttcaaatgat
     5461 tcacagttaa tttgcgacaa tataatttta ttttcacata aactagacgc cttgtcgtct
     5521 tcttcttcgt attccttctc tttttcattt ttctcctcat aaaaattaac atagttatta
     5581 tcgtatccat atatgtatct atcgtataga gtaaattttt tgttgtcata aatatatatg
     5641 tcttttttaa tggggtgtat agtaccgctg cgcatagttt ttctgtaatt tacaacagtg
     5701 ctattttctg gtagttcttc ggagtgtgtt gctttaatta ttaaatttat ataatcaatg
     5761 aatttgggat cgtcggtttt gtacaatatg ttgccggcat agtacgcagc ttcttctagt
     5821 tcaattacac cattttttag cagcaccgga ttaacataac tttccaaaat gttgtacgaa
     5881 ccgttaaaca aaaacagttc acctcccttt tctatactat tgtctgcgag cagttgtttg
     5941 ttgttaaaaa taacagccat tgtaatgaga cgcacaaact aatatcacaa gctgccacca
     6001 ttttgttcgc ccaggactct agctatagtt ctagtggttg gctacgtata ctccggaata
     6061 ttaatagatc atggagataa ttaaaatgat aaccatctcg caaataaata agtattttac
     6121 tgttttcgta acagttttgt aataaaaaaa cctataaat
//
