LOCUS       sequence-224675-        7312 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal HIS6-MBP 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 7312)
  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 7312)
  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: pOCC231
FEATURES             Location/Qualifiers
     source          1..7312
                     /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"
     CDS             230..247
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             260..1360
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1334..1357
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1415..1438
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1547..1852
                     /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     1731..1750
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1908..1928
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(1929..1931,1932..1934,1935..2648)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELY*"
     primer_bind     complement(1974..1995)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2235..2254)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2582..2603
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2778..2912
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2815..2834)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2869..2888
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2985..3357
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3709..3726)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3819..4407)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3899..3918)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4578..5369,5370..5438))
                     /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     5201..5220
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5440..5544)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5845..5867
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5859..5876
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5860..5876
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5887..5906
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5887..5905
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6516..6640
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6678..7283)
                     /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 ggcagcagcc atcaccatca
      241 ccatcacagc agcggcagaa tgaaaatcga agaaggtaaa ctggtaatct ggattaacgg
      301 cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat
      361 taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac
      421 tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc
      481 tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac
      541 ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt
      601 atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc
      661 ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga
      721 accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa
      781 cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac
      841 cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc
      901 agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc
      961 caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca
     1021 accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa
     1081 agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt
     1141 taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggtgaa
     1201 agatccgcgt attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat
     1261 cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg
     1321 tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa
     1381 caacaataac aataacaaca acagcagcgg cagactggaa gttctgttcc aggggccggc
     1441 ggccgcaagc gctatggcca caacctgtta cattgcacaa gataaaaata tatcatcacg
     1501 aacagtaaaa ctgtctgctt acataaacag taatacaagg ggtgttatgc agtttaaggt
     1561 ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga gtgatattat
     1621 tgacacgccc gggcgacgga tggtgatccc cctggccagt gcccgtctgc tgtcagataa
     1681 agtctcccgt gaactttacc cggtggtgca tatcggggat gaaagctggc gcatgatgac
     1741 caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg atctcagcca
     1801 ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat aaatgtcagg
     1861 ctcccttata cacagccagt ggcgcgccgg gatccagcag cggcagagaa aacttgtatt
     1921 tccagggcat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1981 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     2041 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     2101 ggcccaccct cgtgaccacc ttcggctacg gcctgatgtg cttcgcccgc taccccgacc
     2161 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     2221 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     2281 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     2341 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     2401 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     2461 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     2521 acaaccacta cctgagctac cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     2581 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     2641 actaatagaa gcttgtcgag aagtactaga ggatcataat cagccatacc acatttgtag
     2701 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     2761 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     2821 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     2881 aactcatcaa tgtatcttat catgtctgga tctgatcact gcttgagcct aggagatccg
     2941 aaccagataa gtgaaagcct gcatccgcag atcttaatta ataaaacacg atacattgtt
     3001 attagtacat ttattaagcg ctagattctg tgcgttgttg atttacagac aattgttgta
     3061 cgtattttaa taattcatta aatttataat ctttagggtg gtatgttaga gcgaaaatca
     3121 aatgattttc agcgtcttta tatctgaatt taaatattaa atcctcaata gatttgtaaa
     3181 ataggtttcg attagtttca aacaagggtt gtttttccga accgatggct ggactatcta
     3241 atggattttc gctcaacgcc acaaaacttg ccaaatcttg tagcagcaat ctagctttgt
     3301 cgatattcgt ttgtgttttg ttttgtaata aaggttcgac gtcgttcaaa atattatgcg
     3361 cttttgtatt tctttcatca ctgtcgttag tgtacaattg actcgacgta aacacgttaa
     3421 ataacgcttg gacatattta acatcgggcg tgttagcttt attaggccga ttatcgtcgt
     3481 cgtcccaacc ctcgtcgtta gaagttgctt ccgaagacga ttttgccata gccacacgac
     3541 gcctattaat tgtgtcggct aacacgtccg cgatcaaatt tgtagttgag ctttttgggc
     3601 ctcatgggcc ttccgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     3661 ttaacatggt catagctgtt tccttgcgta ttgggcgctc tccgcttcct cgctcactga
     3721 ctcgctgcgc tcggtcgttc gggtaaagcc tggggtgcct aatgagcaaa aggccagcaa
     3781 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     3841 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     3901 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     3961 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4021 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4081 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4141 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4201 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     4261 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4321 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4381 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     4441 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     4501 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     4561 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     4621 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     4681 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     4741 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     4801 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     4861 gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg
     4921 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     4981 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5041 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5101 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5161 atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc
     5221 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5281 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5341 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     5401 aagggaataa gggcgacacg gaaatgttga atactcatta ctcttccttt ttcaatatta
     5461 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     5521 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc
     5581 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     5641 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     5701 ggccgctaca gggcgctccc attcgccatt caggctgcgc aactgttggg aagggcgttt
     5761 cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat
     5821 taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgagc
     5881 gcgacgtaat acgactcact atagggcgaa ttggcggaag gccgtcaagg ccgtgacgtt
     5941 aaaactatta agccatccaa tcgaccgtta gtcgaatcag gaccgctggt gcgagaagcc
     6001 gcgaagtatg gcgaatgcat cgtataacgt gtggagtccg ctcattagag cgtcatgttt
     6061 agacaagaaa gctacatatt taattgatcc cgatgatttt attgataaat tgaccctaac
     6121 tccatacacg gtattctaca atggcggggt tttggtcaaa atttccggac tgcgattgta
     6181 catgctgtta acggctccgc ccactattaa tgaaattaaa aattccaatt ttaaaaaacg
     6241 cagcaagaga aacatttgta tgaaagaatg cgtagaagga aagaaaaatg tcgtggacat
     6301 gctgaacaac aagattaata tgcctccgtg tataaaaaaa atattgaacg atttgaaaga
     6361 aaacaatgta ccgcgcggcg gtatgtacag gaagaggttt atactaaact gttacattgc
     6421 aaacgtggtt tcgtgtgcca agtgtgaaaa ccgatgttta atcaaggctc tgacgcattt
     6481 ctacaaccac gactccaagt gtgtgggtga agtcatgcat cttttaatca aatcccaaga
     6541 tgtgtataaa ccaccaaact gccaaaaaat gaaaactgtg gacaagctct gtccgtttgc
     6601 tggcaactgc aagggtctca atcctatttg taattattga ataataaaac aattataaat
     6661 gctaaatttg ttttttatta acgatacaaa ccaaacgcaa caagaacatt tgtagtatta
     6721 tctataattg aaaacgcgta gttataatcg ctgaggtaat atttaaaatc attttcaaat
     6781 gattcacagt taatttgcga caatataatt ttattttcac ataaactaga cgccttgtcg
     6841 tcttcttctt cgtattcctt ctctttttca tttttctcct cataaaaatt aacatagtta
     6901 ttatcgtatc catatatgta tctatcgtat agagtaaatt ttttgttgtc ataaatatat
     6961 atgtcttttt taatggggtg tatagtaccg ctgcgcatag tttttctgta atttacaaca
     7021 gtgctatttt ctggtagttc ttcggagtgt gttgctttaa ttattaaatt tatataatca
     7081 atgaatttgg gatcgtcggt tttgtacaat atgttgccgg catagtacgc agcttcttct
     7141 agttcaatta caccattttt tagcagcacc ggattaacat aactttccaa aatgttgtac
     7201 gaaccgttaa acaaaaacag ttcacctccc ttttctatac tattgtctgc gagcagttgt
     7261 ttgttgttaa aaataacagc cattgtaatg agacgcacaa actaatatca ca
//
