LOCUS       sequence-320679-        6084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses BFLF1 in mammalian cells with an N-terminal TwinStrep tag 
            and HRV 3C protease cleavage site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6084)
  AUTHORS   Didychuk AL, Gates SN, Gardner MR, Strong LM, Martin A, Glaunsinger 
            BA
  TITLE     A pentameric protein ring with novel architecture is required for 
            herpesviral packaging.
  JOURNAL   Elife. 2021 Feb 8;10. pii: 62261. doi: 10.7554/eLife.62261.
  PUBMED    33554858
REFERENCE   2  (bases 1 to 6084)
  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: "Elife."; 
            date: "2021-02-8"; volume: "10. pii: 62261. doi"; pages: " 
            10.7554/eLife.62261"
COMMENT     Sequence Label: pUE1-TSP-BFLF1
FEATURES             Location/Qualifiers
     source          1..6084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        60..363
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        364..567
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     518..538
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             join(814..819,820..843,844..879,880..903)
                     /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             910..933
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     misc_feature    2413..2985
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2580..2597)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     regulatory      2987..2996
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3267..3286)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3311..3358
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     complement(3445..3462)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3445..3461)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3454..3476)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3670..3689)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3789..3811
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3849..3867)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3935..4039
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4040..4108,4109..4900)
                     /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     complement(4258..4277)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5071..5659
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5560..5579
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5813..5830
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5947..5968
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5983..5988,5989..6006,6007..6013)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6021..6037
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6026..6048
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6045..6061
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6045..6061
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gaattctaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
       61 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
      121 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      181 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      241 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
      301 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
      361 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
      421 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
      481 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
      541 cggtgggagg tctatataag cagagctggt ttagtgaacc gaattcgtac ttcaagaact
      601 gctgatatcg agcttgctac aagggacttt ccgctgggga ctttccaggg aggcgtggcc
      661 tgggcgggac tggggagtgg cgagccctca gatcctgcat ataagcagct gctttttgcc
      721 tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg
      781 aacccactgc ttaagcctcc ccgggctcga atgagtgcgt ggagtcatcc tcaattcgag
      841 aaaggtggag gttctggcgg tggatcggga ggttcagcgt ggagccaccc gcagttcgaa
      901 aaatccggac tggaagttct gttccagggg cccatggagg actttgtgcc ctggaccgtg
      961 gacaatctga aaagccagtt cgaggcagtc ggccttttga tggcccactc ctatcttcca
     1021 gccaacgcgg aagaaggcat agcctaccca ccactcgtcc acacctacga gtccctctct
     1081 ccagccagca cctgccgagt ttgtgatctg ctcgacaccc tggtcaatca ctcagatgcc
     1141 cccgtggcct tctttgagga ctatgccctg ctctgctact actgtctaaa tgccccacga
     1201 gcctggatct ctagcctcat cacgggcatg gactttttac acatcctgat aaaatacttc
     1261 cccatggccg gggggttgga ttccctcttc atgccctcca ggattctggc cattgacatc
     1321 cagcttcact tttacatttg ccggtgcttc ctccccgtgt cctcttctga catgattaga
     1381 aatgccaacc tcggctacta caaattggag tttctgaaat ccatcctcac cggccagtcc
     1441 ccggccaact tctgcttcaa gtccatgtgg cccagaacca cccccacttt tcttaccctc
     1501 cctggaccca gaacctgcaa agattcccag gatgtcccag gggacatcgg caggggccag
     1561 tacaccgccc tgtgctgcca cctgcctacc agaaaccgag ttcagcatcc atttctcagg
     1621 gccgaaaagg gaggactctc tcctgaaatt acaaccaagg ccgactactg tggactacta
     1681 ttaggtacgt ggcaggggac ggacctgctc ggggggccag gccaccatgc tatcggttta
     1741 aacgcggaat acagtggcga tgagctggcg gagttggccc tggccatcac cagaccggag
     1801 gctggcgacc attcgcaggg cccttgcctc ctggccccca tgtttggact aaggcacaag
     1861 aatgcctcac ggaccatctg cccactctgc gagagcctcg gggcccaccc ggacgccaag
     1921 gacacccttg accgcttcaa gagcctgatt ctggactcat tcggcaacaa catcaaaatc
     1981 ctagacagga tcgtgttcct catcaagacc caaaacacgc tgctggatgt gccatgcccc
     2041 aggctaaggg cctggctgca gatgtgcacc ccgcaagact tccacaagca cctgttctgc
     2101 gaccccctgt gtgccatcaa ccacagcatc acgaatccga gcgtgctgtt tggtcagatt
     2161 tacccaccca gcttccaggc ctttaaggca gcgcttgcag ccggtcagaa tctggaacag
     2221 ggggtctgtg actcgttaat tacgcttgtg tacatattta aatccacaca agtggccaga
     2281 gtgggcaaaa caatcctcgt ggatgtcact aaggaactgg acgtggtcct gcgcatccac
     2341 ggccttgacc tggtacagtc ctatcaaact tcccaggtct acgtgtgatc gacctgcagg
     2401 catgcaagct tgcccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga
     2461 ataaggccgg tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa
     2521 tgtgagggcc cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc
     2581 tctcgccaaa ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc
     2641 ttcttgaaga caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg
     2701 cgacaggtgc ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca
     2761 accccagtgc cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tcacctcaag
     2821 cgtattcaac aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct
     2881 ggggcctcgg tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaacg tctaggcccc
     2941 ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataatgcca ccatggagcc
     3001 agtagatcct agactagagc cctggaagca tccaggaagt cagcctaaaa ctgcttgtac
     3061 caattgctat tgtaaaaagt gttgctttca ttgccaagtt tgtttcataa caaaagcctt
     3121 aggcatctcc tatggcagga agaagcggag acagcgacga agagctcatc agaacagtca
     3181 gactcatcaa gcttctctat caaagcaacc cacctcccaa ccccgagggg acccgacagg
     3241 cccgaaggaa taggggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     3301 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     3361 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     3421 ggagcttgat atcgatagct tggcactggc cgtcgtttta caacgtcgtg actgggaaaa
     3481 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     3541 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     3601 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatatggtg
     3661 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     3721 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     3781 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     3841 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     3901 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     3961 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     4021 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     4081 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     4141 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     4201 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     4261 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     4321 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     4381 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     4441 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     4501 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     4561 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     4621 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cagataaagt
     4681 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     4741 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     4801 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     4861 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     4921 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     4981 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     5041 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     5101 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     5161 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     5221 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     5281 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     5341 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     5401 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     5461 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     5521 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     5581 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     5641 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     5701 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     5761 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     5821 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     5881 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     5941 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     6001 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     6061 ccatgattac gaattatcga tatc
//
