LOCUS       sequence-232325-        5454 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  shuttle vector for baculovirus production, using FlashBac bacmid; 
            for recombinant protein with C-terminal TwinStrep, cleavable with 
            3C.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5454)
  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 5454)
  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: pOCC154
FEATURES             Location/Qualifiers
     source          1..5454
                     /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"
     regulatory      46..55
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             159..464
                     /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     343..362
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             502..525
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(526..531,532..555,556..591,592..615)
                     /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"
     polyA_signal    751..885
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(788..807)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     842..861
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     3'UTR           958..1330
                     /label=polyhedrin 3' UTR
                     /note="3' UTR from the Autographa californica 
                     nucleopolyhedrovirus polyhedrin gene (Suzuki et al., 2006)"
     primer_bind     complement(1682..1699)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1792..2380)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1872..1891)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2551..3342,3343..3411))
                     /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     3174..3193
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3412..3516)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3817..3839
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3831..3848
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3832..3848
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3859..3878
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3859..3877
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             4488..4612
                     /codon_start=3
                     /product="baculovirus late expression factor 2"
                     /label=lef2
                     /translation="HLLIKSQDVYKPPNCQKMKTVDKLCPFAGNCKGLNPICNY"
     CDS             complement(4650..5255)
                     /codon_start=1
                     /product="baculovirus ORF603 protein"
                     /label=ORF603
                     /translation="MAVIFNNKQLLADNSIEKGGELFLFNGSYNILESYVNPVLLKNGV
                     IELEEAAYYAGNILYKTDDPKFIDYINLIIKATHSEELPENSTVVNYRKTMRSGTIHPI
                     KKDIYIYDNKKFTLYDRYIYGYDNNYVNFYEEKNEKEKEYEEEDDKASSLCENKIILSQ
                     INCESFENDFKYYLSDYNYAFSIIDNTTNVLVAFGLYR"
     promoter        5363..5454
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     5379..5396
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
ORIGIN
        1 attccggatt attcataccg tcccaccatc gggcgcggat cgccaccatg gcggccgcaa
       61 gcgctatggc cacaacctgt tacattgcac aagataaaaa tatatcatca cgaacagtaa
      121 aactgtctgc ttacataaac agtaatacaa ggggtgttat gcagtttaag gtttacacct
      181 ataaaagaga gagccgttat cgtctgtttg tggatgtaca gagtgatatt attgacacgc
      241 ccgggcgacg gatggtgatc cccctggcca gtgcccgtct gctgtcagat aaagtctccc
      301 gtgaacttta cccggtggtg catatcgggg atgaaagctg gcgcatgatg accaccgata
      361 tggccagtgt gccggtctcc gttatcgggg aagaagtggc tgatctcagc caccgcgaaa
      421 atgacatcaa aaacgccatt aacctgatgt tctggggaat ataaatgtca ggctccctta
      481 tacacagcca gtggcgcgcc gctggaagtt ctgttccagg ggccgagcgc ttggagccat
      541 ccgcagtttg aaaaaggtgg tggtagcggt ggtggttcag gtggtagtgc atggtcacac
      601 cctcagtttg agaaataata gaagcttgtc gagaagtact agaggatcat aatcagccat
      661 accacatttg tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg
      721 aaacataaaa tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac
      781 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
      841 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggatctgatc actgcttgag
      901 cctaggagat ccgaaccaga taagtgaaag cctgcatccg cagatcttaa ttaataaaac
      961 acgatacatt gttattagta catttattaa gcgctagatt ctgtgcgttg ttgatttaca
     1021 gacaattgtt gtacgtattt taataattca ttaaatttat aatctttagg gtggtatgtt
     1081 agagcgaaaa tcaaatgatt ttcagcgtct ttatatctga atttaaatat taaatcctca
     1141 atagatttgt aaaataggtt tcgattagtt tcaaacaagg gttgtttttc cgaaccgatg
     1201 gctggactat ctaatggatt ttcgctcaac gccacaaaac ttgccaaatc ttgtagcagc
     1261 aatctagctt tgtcgatatt cgtttgtgtt ttgttttgta ataaaggttc gacgtcgttc
     1321 aaaatattat gcgcttttgt atttctttca tcactgtcgt tagtgtacaa ttgactcgac
     1381 gtaaacacgt taaataacgc ttggacatat ttaacatcgg gcgtgttagc tttattaggc
     1441 cgattatcgt cgtcgtccca accctcgtcg ttagaagttg cttccgaaga cgattttgcc
     1501 atagccacac gacgcctatt aattgtgtcg gctaacacgt ccgcgatcaa atttgtagtt
     1561 gagctttttg ggcctcatgg gccttccgct cactgcccgc tttccagtcg ggaaacctgt
     1621 cgtgccagct gcattaacat ggtcatagct gtttccttgc gtattgggcg ctctccgctt
     1681 cctcgctcac tgactcgctg cgctcggtcg ttcgggtaaa gcctggggtg cctaatgagc
     1741 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     1801 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     1861 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     1921 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     1981 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     2041 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     2101 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     2161 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     2221 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     2281 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     2341 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     2401 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     2461 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     2521 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     2581 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     2641 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagaaccacg
     2701 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     2761 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     2821 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     2881 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     2941 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3001 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3061 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     3121 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     3181 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     3241 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     3301 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     3361 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     3421 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     3481 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     3541 taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca
     3601 ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag
     3661 atagggttga gtggccgcta cagggcgctc ccattcgcca ttcaggctgc gcaactgttg
     3721 ggaagggcgt ttcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     3781 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     3841 cggccagtga gcgcgacgta atacgactca ctatagggcg aattggcgga aggccgtcaa
     3901 ggccgtgacg ttaaaactat taagccatcc aatcgaccgt tagtcgaatc aggaccgctg
     3961 gtgcgagaag ccgcgaagta tggcgaatgc atcgtataac gtgtggagtc cgctcattag
     4021 agcgtcatgt ttagacaaga aagctacata tttaattgat cccgatgatt ttattgataa
     4081 attgacccta actccataca cggtattcta caatggcggg gttttggtca aaatttccgg
     4141 actgcgattg tacatgctgt taacggctcc gcccactatt aatgaaatta aaaattccaa
     4201 ttttaaaaaa cgcagcaaga gaaacatttg tatgaaagaa tgcgtagaag gaaagaaaaa
     4261 tgtcgtggac atgctgaaca acaagattaa tatgcctccg tgtataaaaa aaatattgaa
     4321 cgatttgaaa gaaaacaatg taccgcgcgg cggtatgtac aggaagaggt ttatactaaa
     4381 ctgttacatt gcaaacgtgg tttcgtgtgc caagtgtgaa aaccgatgtt taatcaaggc
     4441 tctgacgcat ttctacaacc acgactccaa gtgtgtgggt gaagtcatgc atcttttaat
     4501 caaatcccaa gatgtgtata aaccaccaaa ctgccaaaaa atgaaaactg tggacaagct
     4561 ctgtccgttt gctggcaact gcaagggtct caatcctatt tgtaattatt gaataataaa
     4621 acaattataa atgctaaatt tgttttttat taacgataca aaccaaacgc aacaagaaca
     4681 tttgtagtat tatctataat tgaaaacgcg tagttataat cgctgaggta atatttaaaa
     4741 tcattttcaa atgattcaca gttaatttgc gacaatataa ttttattttc acataaacta
     4801 gacgccttgt cgtcttcttc ttcgtattcc ttctcttttt catttttctc ctcataaaaa
     4861 ttaacatagt tattatcgta tccatatatg tatctatcgt atagagtaaa ttttttgttg
     4921 tcataaatat atatgtcttt tttaatgggg tgtatagtac cgctgcgcat agtttttctg
     4981 taatttacaa cagtgctatt ttctggtagt tcttcggagt gtgttgcttt aattattaaa
     5041 tttatataat caatgaattt gggatcgtcg gttttgtaca atatgttgcc ggcatagtac
     5101 gcagcttctt ctagttcaat tacaccattt tttagcagca ccggattaac ataactttcc
     5161 aaaatgttgt acgaaccgtt aaacaaaaac agttcacctc ccttttctat actattgtct
     5221 gcgagcagtt gtttgttgtt aaaaataaca gccattgtaa tgagacgcac aaactaatat
     5281 cacaagctgc caccattttg ttcgcccagg actctagcta tagttctagt ggttggctac
     5341 gtatactccg gaatattaat agatcatgga gataattaaa atgataacca tctcgcaaat
     5401 aaataagtat tttactgttt tcgtaacagt tttgtaataa aaaaacctat aaat
//
