LOCUS       sequence-224688-        6682 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for rec. protein with N-terminal TwinStrep-MBP tag, cleavable with 
            3C and C-terminal HIS6, cleavable with 3C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6682)
  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 6682)
  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: pOCC305
FEATURES             Location/Qualifiers
     source          1..6682
                     /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             join(224..229,230..253,254..289,290..313)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="SAWSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     CDS             326..1426
                     /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     1400..1423
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1481..1504
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1613..1918
                     /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     1797..1816
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1956..1979
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1989..2006
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2148..2282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2185..2204)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2239..2258
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           2355..2727
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3079..3096)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3189..3777)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3269..3288)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3948..4739,4740..4808))
                     /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     4571..4590
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4810..4914)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5215..5237
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5229..5246
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5230..5246
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5257..5276
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5257..5275
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5886..6010
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6048..6653)
                     /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 ggcagcgctt ggagccatcc
      241 gcagtttgaa aaaggtggtg gtagcggtgg tggttcaggt ggtagtgcat ggtcacaccc
      301 tcagtttgag aaaagcagcg gcagaatgaa aatcgaagaa ggtaaactgg taatctggat
      361 taacggcgat aaaggctata acggtctcgc tgaagtcggt aagaaattcg agaaagatac
      421 cggaattaaa gtcaccgttg agcatccgga taaactggaa gagaaattcc cacaggttgc
      481 ggcaactggc gatggccctg acattatctt ctgggcacac gaccgctttg gtggctacgc
      541 tcaatctggc ctgttggctg aaatcacccc ggacaaagcg ttccaggaca agctgtatcc
      601 gtttacctgg gatgccgtac gttacaacgg caagctgatt gcttacccga tcgctgttga
      661 agcgttatcg ctgatttata acaaagatct gctgccgaac ccgccaaaaa cctgggaaga
      721 gatcccggcg ctggataaag aactgaaagc gaaaggtaag agcgcgctga tgttcaacct
      781 gcaagaaccg tacttcacct ggccgctgat tgctgctgac gggggttatg cgttcaagta
      841 tgaaaacggc aagtacgaca ttaaagacgt gggcgtggat aacgctggcg cgaaagcggg
      901 tctgaccttc ctggttgacc tgattaaaaa caaacacatg aatgcagaca ccgattactc
      961 catcgcagaa gctgccttta ataaaggcga aacagcgatg accatcaacg gcccgtgggc
     1021 atggtccaac atcgacacca gcaaagtgaa ttatggtgta acggtactgc cgaccttcaa
     1081 gggtcaacca tccaaaccgt tcgttggcgt gctgagcgca ggtattaacg ccgccagtcc
     1141 gaacaaagag ctggcaaaag agttcctcga aaactatctg ctgactgatg aaggtctgga
     1201 agcggttaat aaagacaaac cgctgggtgc cgtagcgctg aagtcttacg aggaagagtt
     1261 ggtgaaagat ccgcgtattg ccgccactat ggaaaacgcc cagaaaggtg aaatcatgcc
     1321 gaacatcccg cagatgtccg ctttctggta tgccgtgcgt actgcggtga tcaacgccgc
     1381 cagcggtcgt cagactgtcg atgaagccct gaaagacgcg cagactaatt cgagctcgaa
     1441 caacaacaac aataacaata acaacaacag cagcggcaga ctggaagttc tgttccaggg
     1501 gccggcggcc gcaagcgcta tggccacaac ctgttacatt gcacaagata aaaatatatc
     1561 atcacgaaca gtaaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgcagtt
     1621 taaggtttac acctataaaa gagagagccg ttatcgtctg tttgtggatg tacagagtga
     1681 tattattgac acgcccgggc gacggatggt gatccccctg gccagtgccc gtctgctgtc
     1741 agataaagtc tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa gctggcgcat
     1801 gatgaccacc gatatggcca gtgtgccggt ctccgttatc ggggaagaag tggctgatct
     1861 cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg atgttctggg gaatataaat
     1921 gtcaggctcc cttatacaca gccagtggcg cgccgctgga agttctgttc caggggccgg
     1981 gcagcagcca tcaccatcac catcacagtg gataatagaa gcttgtcgag aagtactaga
     2041 ggatcataat cagccatacc acatttgtag aggttttact tgctttaaaa aacctcccac
     2101 acctccccct gaacctgaaa cataaaatga atgcaattgt tgttgttaac ttgtttattg
     2161 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     2221 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgga
     2281 tctgatcact gcttgagcct aggagatccg aaccagataa gtgaaagcct gcatccgcag
     2341 atcttaatta ataaaacacg atacattgtt attagtacat ttattaagcg ctagattctg
     2401 tgcgttgttg atttacagac aattgttgta cgtattttaa taattcatta aatttataat
     2461 ctttagggtg gtatgttaga gcgaaaatca aatgattttc agcgtcttta tatctgaatt
     2521 taaatattaa atcctcaata gatttgtaaa ataggtttcg attagtttca aacaagggtt
     2581 gtttttccga accgatggct ggactatcta atggattttc gctcaacgcc acaaaacttg
     2641 ccaaatcttg tagcagcaat ctagctttgt cgatattcgt ttgtgttttg ttttgtaata
     2701 aaggttcgac gtcgttcaaa atattatgcg cttttgtatt tctttcatca ctgtcgttag
     2761 tgtacaattg actcgacgta aacacgttaa ataacgcttg gacatattta acatcgggcg
     2821 tgttagcttt attaggccga ttatcgtcgt cgtcccaacc ctcgtcgtta gaagttgctt
     2881 ccgaagacga ttttgccata gccacacgac gcctattaat tgtgtcggct aacacgtccg
     2941 cgatcaaatt tgtagttgag ctttttgggc ctcatgggcc ttccgctcac tgcccgcttt
     3001 ccagtcggga aacctgtcgt gccagctgca ttaacatggt catagctgtt tccttgcgta
     3061 ttgggcgctc tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc gggtaaagcc
     3121 tggggtgcct aatgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     3181 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     3241 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     3301 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     3361 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     3421 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3481 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     3541 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3601 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
     3661 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3721 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     3781 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3841 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3901 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3961 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     4021 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     4081 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     4141 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     4201 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     4261 gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     4321 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     4381 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     4441 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     4501 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     4561 tcaacacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     4621 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     4681 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     4741 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     4801 atactcatta ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     4861 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     4921 tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     4981 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     5041 tcaaaagaat agaccgagat agggttgagt ggccgctaca gggcgctccc attcgccatt
     5101 caggctgcgc aactgttggg aagggcgttt cggtgcgggc ctcttcgcta ttacgccagc
     5161 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
     5221 cacgacgttg taaaacgacg gccagtgagc gcgacgtaat acgactcact atagggcgaa
     5281 ttggcggaag gccgtcaagg ccgtgacgtt aaaactatta agccatccaa tcgaccgtta
     5341 gtcgaatcag gaccgctggt gcgagaagcc gcgaagtatg gcgaatgcat cgtataacgt
     5401 gtggagtccg ctcattagag cgtcatgttt agacaagaaa gctacatatt taattgatcc
     5461 cgatgatttt attgataaat tgaccctaac tccatacacg gtattctaca atggcggggt
     5521 tttggtcaaa atttccggac tgcgattgta catgctgtta acggctccgc ccactattaa
     5581 tgaaattaaa aattccaatt ttaaaaaacg cagcaagaga aacatttgta tgaaagaatg
     5641 cgtagaagga aagaaaaatg tcgtggacat gctgaacaac aagattaata tgcctccgtg
     5701 tataaaaaaa atattgaacg atttgaaaga aaacaatgta ccgcgcggcg gtatgtacag
     5761 gaagaggttt atactaaact gttacattgc aaacgtggtt tcgtgtgcca agtgtgaaaa
     5821 ccgatgttta atcaaggctc tgacgcattt ctacaaccac gactccaagt gtgtgggtga
     5881 agtcatgcat cttttaatca aatcccaaga tgtgtataaa ccaccaaact gccaaaaaat
     5941 gaaaactgtg gacaagctct gtccgtttgc tggcaactgc aagggtctca atcctatttg
     6001 taattattga ataataaaac aattataaat gctaaatttg ttttttatta acgatacaaa
     6061 ccaaacgcaa caagaacatt tgtagtatta tctataattg aaaacgcgta gttataatcg
     6121 ctgaggtaat atttaaaatc attttcaaat gattcacagt taatttgcga caatataatt
     6181 ttattttcac ataaactaga cgccttgtcg tcttcttctt cgtattcctt ctctttttca
     6241 tttttctcct cataaaaatt aacatagtta ttatcgtatc catatatgta tctatcgtat
     6301 agagtaaatt ttttgttgtc ataaatatat atgtcttttt taatggggtg tatagtaccg
     6361 ctgcgcatag tttttctgta atttacaaca gtgctatttt ctggtagttc ttcggagtgt
     6421 gttgctttaa ttattaaatt tatataatca atgaatttgg gatcgtcggt tttgtacaat
     6481 atgttgccgg catagtacgc agcttcttct agttcaatta caccattttt tagcagcacc
     6541 ggattaacat aactttccaa aatgttgtac gaaccgttaa acaaaaacag ttcacctccc
     6601 ttttctatac tattgtctgc gagcagttgt ttgttgttaa aaataacagc cattgtaatg
     6661 agacgcacaa actaatatca ca
//
