LOCUS       sequence-373430-        7687 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses C terminal 3xFLAG tagged PB1 subunit of Influenza 
            B/Brisbane/60/2008 virus polymerase in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7687)
  AUTHORS   Kedia N, Banerjee S, Mondal A
  TITLE     A Comprehensive Roadmap Towards the Generation of an Influenza B 
            Reporter Assay Using a Single DNA Polymerase-Based Cloning of the 
            Reporter RNA Construct.
  JOURNAL   Front Microbiol. 2022 May 25;13:868367. doi: 
            10.3389/fmicb.2022.868367. eCollection 2022.
  PUBMED    35694292
REFERENCE   2  (bases 1 to 7687)
  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: "Front 
            Microbiol."; date: "2022-05-25"; volume: "13:868367. doi"; pages: " 
            10.3389/fmicb.2022.868367. eCollection 2022"
COMMENT     Sequence Label: pcDNA3-PB1-3x-FLAG-B/BRISBANE/60/2008
FEATURES             Location/Qualifiers
     source          1..7687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3212..3277
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     promoter        complement(3312..3330)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3313..3330)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3350..3367)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3356..3580
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3626..4054
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3713..3732)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3923..3944
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4063..4083)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4107..4317
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        4129..4325
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4176..4311
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4238..4257
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4392..5186
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4446..4465)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5056..5075
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5360..5481
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5397..5416)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5451..5470
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5530..5546)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5530..5546)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5543..5565)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5554..5570
                     /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)."
     promoter        complement(join(5578..5584,5585..5602,5603..5608))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5623..5644
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5761..5778)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5932..6520)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6012..6031)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6691..7482,7483..7551))
                     /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     7314..7333
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7552..7656)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat ccactagtaa cggccgccag tgtgctggaa ttcaccatga atataaatcc
      961 ttattttctc ttcatagatg tgcccgtaca ggcagcaatt tcaacaacat tcccatacac
     1021 tggtgttccc ccttattccc atggaacagg aacaggctac acaatagaca ccgtgatcag
     1081 aacgcatgag tactcaaaca aggggaaaca gtacatttct gatgttacag gatgcacaat
     1141 ggtagatcca acaaatggac cattacccga agataatgag ccgagtgcct atgcgcaatt
     1201 agattgcgtt ttagaggctt tggatagaat ggatgaagaa cacccaggtc tttttcaagc
     1261 agcctcacag aatgctatgg aggccctaat ggtcacaact gtagacaaat taacccaggg
     1321 gagacagact tttgattgga cagtatgcag aaaccaacct gctgcaacgg cactgaacac
     1381 aacaataacc tcttttaggt tgaatgattt aaatggagcc gacaaaggtg gattaatacc
     1441 tttttgccag gatatcattg attcattaga ccgacctgaa atgactttct tctcagtaaa
     1501 gaatataaag aaaaaattgc ctgccaaaaa cagaaagggt ttcctcataa agaggatacc
     1561 aatgaaggta aaagacaaaa taaccaaagt ggaatacatc aaaagagcat tatcattaaa
     1621 cacaatgaca aaagacgctg aaagaggcaa actgaaaaga agagcgattg ccactgctgg
     1681 aatacaaatc agagggtttg tattagtagt tgaaaacttg gctaaaaata tatgtgaaaa
     1741 tctagaacaa agtggtttac cagtaggtgg aaacgagaag aaagccaaac tgtcaaacgc
     1801 agtggccaaa atgctcagta actgcccacc aggagggatt agcatgacag taacaggaga
     1861 caatacaaaa tggaatgaat gtttaaaccc aagaatcttt ttggctatga ctgaaagaat
     1921 aaccagagac agcccagttt ggttcaggga tttttgtagt atagcaccgg tcctgttctc
     1981 caataagata gcaagattgg ggaaagggtt tatgataaca agcaaaacaa aaagactgaa
     2041 ggctcaaata ccttgtcctg atctgtttag tataccgtta gaaagatata atgaagaaac
     2101 aagggcaaaa ttgaaaaagc taaaaccatt cttcaatgaa gaaggaactg catctttgtc
     2161 gcctgggatg atgatgggaa tgtttaatat gctatctacc gtgttgggag tagctgcact
     2221 aggtatcaag aacattggaa acaaagaata cttatgggat ggactgcaat cttctgatga
     2281 ttttgctctg tttgttaatg caaaggatga agaaacatgt atggaaggaa taaacgactt
     2341 ttaccgaaca tgtaaattat tgggagtaaa catgagcaaa aagaaaagtt actgtaatga
     2401 gactggaatg tttgaattta caagcatgtt ctacagagat ggatttgtat ctaattttgc
     2461 aatggaactc ccttcgtttg gggttgctgg agtaaatgaa tcagcagata tggcaatagg
     2521 aatgacaata ataaagaaca acatgatcaa caatggaatg ggtccggcaa cagcacaaac
     2581 agccatacag ttattcatag ctgattatag atacacctac aaatgccaca ggggagattc
     2641 caaagtagaa ggaaagagaa tgaaaatcat aaaggagtta tgggaaaaca ctaaaggaag
     2701 agatggtcta ttagtagcag atggtgggcc caacatttac aatttgagaa acctgcatat
     2761 cccagaaata gtattaaagt ataatctaat ggaccctgaa tacaaagggc ggttacttca
     2821 tcctcaaaat ccctttgtgg gacatttgtc tattgagggc atcaaagagg cagacataac
     2881 cccagcacat ggtccagtaa agaaaatgga ctacgatgcg gtgtctggaa ctcatagttg
     2941 gagaaccaaa agaaacagat ctatactaaa cactgatcag aggaacatga ttcttgagga
     3001 acaatgctac gctaaatgtt gcaacctatt tgaggcctgt tttaacagtg catcatacag
     3061 gaagccagtg ggtcaacata gcatgcttga ggctatggcc cacagattaa gaatggatgc
     3121 acgattagat tatgaatcag ggagaatgtc aaaggatgat tttgagaaag caatggctca
     3181 ccttggtgag attgggtaca tagcggccgc cgactacaaa gaccatgacg gtgattataa
     3241 agatcatgac atcgactaca aggatgacga tgacaagtag gccgctcgag catgcatcta
     3301 gagggcccta ttctatagtg tcacctaaat gctagagctc gctgatcagc ctcgactgtg
     3361 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     3421 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     3481 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     3541 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc
     3601 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt
     3661 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     3721 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     3781 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     3841 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     3901 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     3961 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     4021 aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag
     4081 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     4141 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     4201 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     4261 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     4321 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     4381 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     4441 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     4501 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     4561 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     4621 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     4681 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     4741 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     4801 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     4861 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     4921 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     4981 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     5041 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     5101 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     5161 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     5221 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     5281 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     5341 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     5401 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     5461 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     5521 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5581 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5641 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5701 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     5761 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     5821 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     5881 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     5941 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     6001 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     6061 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     6121 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     6181 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     6241 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     6301 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     6361 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     6421 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     6481 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     6541 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     6601 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     6661 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     6721 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     6781 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     6841 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     6901 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     6961 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     7021 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     7081 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     7141 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     7201 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     7261 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     7321 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     7381 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     7441 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     7501 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     7561 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     7621 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     7681 tgacgtc
//
