LOCUS       sequence-224682-        7327 bp    DNA     circular SYN 15-APR-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with N-terminal MBP tag, cleavable with 3C, 
            and C-terminal monomeric GFP and HIS6, cleavable with TEV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7327)
  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 7327)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pOCC120
FEATURES             Location/Qualifiers
     source          1..7327
                     /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             224..1324
                     /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     1298..1321
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1379..1402
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1511..1816
                     /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     1695..1714
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             join(1884..1886,1887..1889,1890..2600)
                     /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(1929..1950)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2190..2209)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2537..2558
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2601..2624
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2634..2651
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    2793..2927
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2830..2849)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2884..2903
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           3000..3372
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(3724..3741)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3834..4422)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3914..3933)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4593..5384,5385..5453))
                     /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     5216..5235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5455..5559)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5860..5882
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5874..5891
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5875..5891
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5902..5921
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5902..5920
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6531..6655
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(6693..7298)
                     /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 ggcatgaaaa tcgaagaagg
      241 taaactggta atctggatta acggcgataa aggctataac ggtctcgctg aagtcggtaa
      301 gaaattcgag aaagataccg gaattaaagt caccgttgag catccggata aactggaaga
      361 gaaattccca caggttgcgg caactggcga tggccctgac attatcttct gggcacacga
      421 ccgctttggt ggctacgctc aatctggcct gttggctgaa atcaccccgg acaaagcgtt
      481 ccaggacaag ctgtatccgt ttacctggga tgccgtacgt tacaacggca agctgattgc
      541 ttacccgatc gctgttgaag cgttatcgct gatttataac aaagatctgc tgccgaaccc
      601 gccaaaaacc tgggaagaga tcccggcgct ggataaagaa ctgaaagcga aaggtaagag
      661 cgcgctgatg ttcaacctgc aagaaccgta cttcacctgg ccgctgattg ctgctgacgg
      721 gggttatgcg ttcaagtatg aaaacggcaa gtacgacatt aaagacgtgg gcgtggataa
      781 cgctggcgcg aaagcgggtc tgaccttcct ggttgacctg attaaaaaca aacacatgaa
      841 tgcagacacc gattactcca tcgcagaagc tgcctttaat aaaggcgaaa cagcgatgac
      901 catcaacggc ccgtgggcat ggtccaacat cgacaccagc aaagtgaatt atggtgtaac
      961 ggtactgccg accttcaagg gtcaaccatc caaaccgttc gttggcgtgc tgagcgcagg
     1021 tattaacgcc gccagtccga acaaagagct ggcaaaagag ttcctcgaaa actatctgct
     1081 gactgatgaa ggtctggaag cggttaataa agacaaaccg ctgggtgccg tagcgctgaa
     1141 gtcttacgag gaagagttgg tgaaagatcc gcgtattgcc gccactatgg aaaacgccca
     1201 gaaaggtgaa atcatgccga acatcccgca gatgtccgct ttctggtatg ccgtgcgtac
     1261 tgcggtgatc aacgccgcca gcggtcgtca gactgtcgat gaagccctga aagacgcgca
     1321 gactaattcg agctcgaaca acaacaacaa taacaataac aacaacagca gcggcagact
     1381 ggaagttctg ttccaggggc cggcggccgc aagcgctatg gccacaacct gttacattgc
     1441 acaagataaa aatatatcat cacgaacagt aaaactgtct gcttacataa acagtaatac
     1501 aaggggtgtt atgcagttta aggtttacac ctataaaaga gagagccgtt atcgtctgtt
     1561 tgtggatgta cagagtgata ttattgacac gcccgggcga cggatggtga tccccctggc
     1621 cagtgcccgt ctgctgtcag ataaagtctc ccgtgaactt tacccggtgg tgcatatcgg
     1681 ggatgaaagc tggcgcatga tgaccaccga tatggccagt gtgccggtct ccgttatcgg
     1741 ggaagaagtg gctgatctca gccaccgcga aaatgacatc aaaaacgcca ttaacctgat
     1801 gttctgggga atataaatgt caggctccct tatacacagc cagtggcgcg ccgggatccg
     1861 ctggctccgc tgctggttct ggcatggtga gcaagggcga ggagctgttc accggggtgg
     1921 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     1981 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca
     2041 agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca
     2101 gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     2161 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     2221 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     2281 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     2341 tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg
     2401 aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     2461 ccgtgctgct gcccgacaac cactacctga gcacccagtc caaactgagc aaagacccca
     2521 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     2581 gcatggacga gctgtacaag ctggaagttc tgttccaggg gccgggcagc agccatcacc
     2641 atcaccatca cagtggataa tagaagcttg tcgagaagta ctagaggatc ataatcagcc
     2701 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
     2761 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     2821 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     2881 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatctga tcactgcttg
     2941 agcctaggag atccgaacca gataagtgaa agcctgcatc cgcagatctt aattaataaa
     3001 acacgataca ttgttattag tacatttatt aagcgctaga ttctgtgcgt tgttgattta
     3061 cagacaattg ttgtacgtat tttaataatt cattaaattt ataatcttta gggtggtatg
     3121 ttagagcgaa aatcaaatga ttttcagcgt ctttatatct gaatttaaat attaaatcct
     3181 caatagattt gtaaaatagg tttcgattag tttcaaacaa gggttgtttt tccgaaccga
     3241 tggctggact atctaatgga ttttcgctca acgccacaaa acttgccaaa tcttgtagca
     3301 gcaatctagc tttgtcgata ttcgtttgtg ttttgttttg taataaaggt tcgacgtcgt
     3361 tcaaaatatt atgcgctttt gtatttcttt catcactgtc gttagtgtac aattgactcg
     3421 acgtaaacac gttaaataac gcttggacat atttaacatc gggcgtgtta gctttattag
     3481 gccgattatc gtcgtcgtcc caaccctcgt cgttagaagt tgcttccgaa gacgattttg
     3541 ccatagccac acgacgccta ttaattgtgt cggctaacac gtccgcgatc aaatttgtag
     3601 ttgagctttt tgggcctcat gggccttccg ctcactgccc gctttccagt cgggaaacct
     3661 gtcgtgccag ctgcattaac atggtcatag ctgtttcctt gcgtattggg cgctctccgc
     3721 ttcctcgctc actgactcgc tgcgctcggt cgttcgggta aagcctgggg tgcctaatga
     3781 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3841 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3901 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3961 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4021 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4081 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4141 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4201 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4261 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4321 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4381 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4441 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4501 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4561 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4621 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4681 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4741 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4801 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4861 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg
     4921 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4981 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5041 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5101 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5161 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaac acgggataat
     5221 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5281 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5341 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5401 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact cattactctt
     5461 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     5521 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     5581 acctaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     5641 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     5701 gagatagggt tgagtggccg ctacagggcg ctcccattcg ccattcaggc tgcgcaactg
     5761 ttgggaaggg cgtttcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     5821 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     5881 cgacggccag tgagcgcgac gtaatacgac tcactatagg gcgaattggc ggaaggccgt
     5941 caaggccgtg acgttaaaac tattaagcca tccaatcgac cgttagtcga atcaggaccg
     6001 ctggtgcgag aagccgcgaa gtatggcgaa tgcatcgtat aacgtgtgga gtccgctcat
     6061 tagagcgtca tgtttagaca agaaagctac atatttaatt gatcccgatg attttattga
     6121 taaattgacc ctaactccat acacggtatt ctacaatggc ggggttttgg tcaaaatttc
     6181 cggactgcga ttgtacatgc tgttaacggc tccgcccact attaatgaaa ttaaaaattc
     6241 caattttaaa aaacgcagca agagaaacat ttgtatgaaa gaatgcgtag aaggaaagaa
     6301 aaatgtcgtg gacatgctga acaacaagat taatatgcct ccgtgtataa aaaaaatatt
     6361 gaacgatttg aaagaaaaca atgtaccgcg cggcggtatg tacaggaaga ggtttatact
     6421 aaactgttac attgcaaacg tggtttcgtg tgccaagtgt gaaaaccgat gtttaatcaa
     6481 ggctctgacg catttctaca accacgactc caagtgtgtg ggtgaagtca tgcatctttt
     6541 aatcaaatcc caagatgtgt ataaaccacc aaactgccaa aaaatgaaaa ctgtggacaa
     6601 gctctgtccg tttgctggca actgcaaggg tctcaatcct atttgtaatt attgaataat
     6661 aaaacaatta taaatgctaa atttgttttt tattaacgat acaaaccaaa cgcaacaaga
     6721 acatttgtag tattatctat aattgaaaac gcgtagttat aatcgctgag gtaatattta
     6781 aaatcatttt caaatgattc acagttaatt tgcgacaata taattttatt ttcacataaa
     6841 ctagacgcct tgtcgtcttc ttcttcgtat tccttctctt tttcattttt ctcctcataa
     6901 aaattaacat agttattatc gtatccatat atgtatctat cgtatagagt aaattttttg
     6961 ttgtcataaa tatatatgtc ttttttaatg gggtgtatag taccgctgcg catagttttt
     7021 ctgtaattta caacagtgct attttctggt agttcttcgg agtgtgttgc tttaattatt
     7081 aaatttatat aatcaatgaa tttgggatcg tcggttttgt acaatatgtt gccggcatag
     7141 tacgcagctt cttctagttc aattacacca ttttttagca gcaccggatt aacataactt
     7201 tccaaaatgt tgtacgaacc gttaaacaaa aacagttcac ctcccttttc tatactattg
     7261 tctgcgagca gttgtttgtt gttaaaaata acagccattg taatgagacg cacaaactaa
     7321 tatcaca
//
