LOCUS       sequence-473499-        8458 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  insect expression of E2F2 for protein purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8458)
  AUTHORS   Clijsters L, Hoencamp C, Calis JJA, Marzio A, Handgraaf SM, Cuitino 
            MC, Rosenberg BR, Leone G, Pagano M
  TITLE     Cyclin F Controls Cell-Cycle Transcriptional Outputs by Directing 
            the Degradation of the Three Activator E2Fs.
  JOURNAL   Mol Cell. 2019 Jun 20;74(6):1264-1277.e7. doi: 
            10.1016/j.molcel.2019.04.010. Epub 2019 May 23.
  PUBMED    31130363
REFERENCE   2  (bases 1 to 8458)
  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"; doi: 
            "10.1016/j.molcel.2019.04.010"; journalName: "Mol Cell"; date: 
            "2019-06-20- 20"; volume: "74"; issue: "6"; pages: "1264-1277.e7"
COMMENT     Sequence Label: NLS-Cherry-PCNA-P2A-Venus-E2F2
FEATURES             Location/Qualifiers
     source          1..8458
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     332..351
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     591..612
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(706..762)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(1600..2304)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1875..1894)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2144..2162
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2341..2361)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(2392..2808)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     complement(2408..2424)
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     complement(2440..2462)
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     misc_feature    3009..3208
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     LTR             3271..3446
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     primer_bind     complement(4048..4066)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4134..4238
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4239..4307,4308..5099)
                     /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     complement(4457..4476)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5270..5858
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5759..5778
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6012..6029
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6044..6184
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6110..6129)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6270..6292
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     LTR             6395..6988
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             complement(join(8399..8458,1..651,652..654))
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(join(8441..8458,1..4))
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 catgtgatcg cgcttctcgt tggggtcttt gctcagggcg gactggtagc tcaggtagtg
       61 gttgtcgggc agcagcacgg ggccgtcgcc gatgggggtg ttctgctggt agtggtcggc
      121 gagctgcacg ccgccgtcct cgatgttgtg gcggatcttg aagttggcct tgatgccgtt
      181 cttctgcttg tcggcggtga tatagacgtt gtggctgttg tagttgtact ccagcttgtg
      241 ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc agctcgatgc ggttcaccag
      301 ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag ttgccgtcgt ccttgaagaa
      361 gatggtgcgc tcctggacgt agccttcggg catggcggac ttgaagaagt cgtgctgctt
      421 catgtggtcg gggtagcggg cgaagcactg taggccgtag cccagagtgg tcacgagggt
      481 gggccagggc acgggcagct tgccggtggt gcagatcagc ttcagggtca gcttgccgta
      541 ggtggcatcg ccctcgccct cgccggacac gctgaacttg tggccgttta cgtcgccgtc
      601 cagctcgacc aggatgggca ccaccccggt gaacagctcc tcgcccttgc tcaccagggt
      661 gcgatcccgg gcccgcggta ccgtcgactg cagaattcga agcttaggtc cagggttctc
      721 ctccacgtct ccagcctgct tcagcaggct gaagttagta gctccgcttc cgagcttgag
      781 ctcgactcga gcagatcctt cttcatcctc gatcttggga gccaagtagt attttaagtg
      841 tcccatatcc gcaattttat actctacaac aaggggtaca tctgcagaca tactgagtgt
      901 caccgttgaa gagagtggag tggcttttgt aaagaagttc aggtacctca gtgcaaaagt
      961 tagttgaact ggttcattca tctctatggt aacagcttcc tcctctttat cgacattact
     1021 tgtctgtgac aatttaatgt ttccatttcc aagttctcca cttgcagaaa atttcactcc
     1081 gtcttttgca caggaaatta caacagcatc tccaatatgg ctgagatctc ggcatatacg
     1141 tgcaaattca ccagaaggca tctttactac acagctgtac tcctgttctg gaattccaag
     1201 ttgttcaaca tctaaatcca tcaacttcat ttcatagtct gaaactttct cctggtttgg
     1261 tgcttcaaat actagcgcca aggtatccgc gttatcttcg gcccttagtg taatgatatc
     1321 ttcattgccg gcgcatttta gtattttgga catactggtg aggttcacgc ccatggccag
     1381 gttgcggtcg cagcggtagg tgtcgaagcc ctcagaccgc agggtgagct gcaccaaaga
     1441 gacgtgggac gagtccatgc tctgcaggtt tacaccgctg gagctaatat cccagcaggc
     1501 ctcgttgatg aggtccttga gtgcctccaa caccttcttg aggatggagc cctggaccag
     1561 gcgcgcctcg aacatttgag ctcgagatct gagtccggac ttgtacagct cgtccatgcc
     1621 gccggtggag tggcggccct cggcgcgttc gtactgttcc acgatggtgt agtcctcgtt
     1681 gtgggaggtg atgtccaact tgatgttgac gttgtaggcg ccgggcagct gcacgggctt
     1741 cttggccttg taggtggtct tgacctcagc gtcgtagtgg ccgccgtcct tcagcttcag
     1801 cctctgcttg atctcgccct tcagggcgcc gtcctcgggg tacatccgct cggaggaggc
     1861 ctcccagccc atggtcttct tctgcattac ggggccgtcg gaggggaagt tggtgccgcg
     1921 cagcttcacc ttgtagatga actcgccgtc ctgcagggag gagtcctggg tcacggtcac
     1981 cacgccgccg tcctcgaagt tcatcacgcg ctcccacttg aagccctcgg ggaaggacag
     2041 cttcaagtag tcggggatgt cggcggggtg cttcacgtag gccttggagc cgtacatgaa
     2101 ctgaggggac aggatgtccc aggcgaaggg cagggggcca cccttggtca ccttcagctt
     2161 ggcggtctgg gtgccctcgt aggggcggcc ctcgccctcg ccctcgatct cgaactcgtg
     2221 gccgttcacg gagccctcca tgtgcacctt gaagcgcatg aactccttga tgatggccat
     2281 gttatcctcc tcgcccttgc tcacggccgc cacagaagct tctcgagatc cagccacggg
     2341 tacctttctc ttcttttttg gtgatgctcc catggtggat ctggcaattc cggcgcctag
     2401 agaaggagtg agggctggat aaagggagga tcgaggcggg gtcgaacgag gaggttcaag
     2461 ggggagagac ggggcggatg gaggaagagg aggcggaggc ttagggtgta caaagggctt
     2521 gacccaggga ggggggtcaa aagccaaggc ttcccaggtc acgatgtagg ggacctggtc
     2581 tgggtgtcca tgcgggccag gtgaaaagac cttgatctta acctgggtga tgaggtctcg
     2641 gttaaaggtg ccgtctcgcg gccatccgac gttaaaggtt ggccattctg cagagcagaa
     2701 ggtaacccaa cgtctcttct tgacatctac cgactggttg tgagcgatcc gctcgacatc
     2761 tttccagtga cctaaggtca aacttaaggg agtggtaaca gtctggccct aattttcaga
     2821 caaatacaga aacacagtca gacagagaca acacagaacg atgctgcagc gctgcagcag
     2881 acaagacgcg cggcttcggt tccaaaccga aagcaaaaat tcagacggag gcgggaactg
     2941 ttttaggttc tcgtctccta ccagaaccac atatcctgac ggggtcggat tccacatcga
     3001 ctcccttcct caggtcgggc cacaaaaacg gcccccaaag tccctgggac gtctcccagg
     3061 gttgcggccg ggtgttcaga actcgtcagt tccaccacgg gtccgccaga tacagagcta
     3121 gttagctaac tagtacagac gcaggcgcat aacatcaaac atagacacta gacaatcgga
     3181 cagacacaga taagttgctg gccagcttac ctcccggtgg tgggtcggtg gtccctgggc
     3241 aggggtctcc aaatcccgga cgagccccca aatgaaagac ccccgtcgtg ggtagtcaat
     3301 cactcagagg agaccctccc aaggaacagc gagaccacga ttcggatgca aacagcaaga
     3361 ggctttattg ggaatacggg tacccgggcg acgcagtcta tcggaagact ggcgcgccga
     3421 gtgaggggtt gtgggctctt ttattgagct cggagagcgg aagcgcgcga acagaagcga
     3481 gaagcgaact gattggttag ttcaaataag gtacagggtc attttcaggt ccttggggca
     3541 ccctggaaac atctgatgat tcactagaaa ctgctgaggg ctggaccgca tctggggacc
     3601 atctgttctt ggccccgagc cggggcagga actgcttacc acagatatcc tgtttggccc
     3661 atcactcagc tgtctcatct gttcttggcc ctgagccggg gcaggaaccg cttaccacag
     3721 atatcctgtt tggtattcag ctgtttcttt gttcctgacc ttgatctgaa cttttctatt
     3781 ctcagttatg tatttttcca tgccttgcaa agtggcgtta cttaagctag cttgccacct
     3841 acgggtgggg tctttcaaaa gcaaagaaac ggaagggccg cggctttggc tccggtgagt
     3901 gtggggaata gggtagagtg gtctaagagg cagaagcccg cgaatttggg agtgtgaggg
     3961 ggacacattt ggaggacagt tttcggtgat ctggtatgaa ttgctagcaa ttgctagcaa
     4021 ttgctagcaa ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     4081 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     4141 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4201 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4261 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4321 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4381 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4441 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc
     4501 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4561 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     4621 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     4681 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     4741 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt
     4801 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     4861 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     4921 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     4981 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5041 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5101 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5161 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5221 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5281 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5341 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5401 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5461 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5521 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     5581 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     5641 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     5701 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     5761 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     5821 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     5881 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     5941 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     6001 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc
     6061 tccttacgca tctgtgcggt atttcacacc gcatatggtg cactctcagt acaatctgct
     6121 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     6181 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     6241 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     6301 gtcatcaccg aaacgcgcga ggcagccagc ttacctcccg gtggtgggtc ggtggtccct
     6361 gggcaggggt ctcccgatcc cggacgagcc cccaaatgaa agacccccgc tgacgggtag
     6421 tcaatcactc agaggagacc ctcccaagga acagcgagac cacaagtcgg atgcaactgc
     6481 aagagggttt attggataca cgggtacccg ggcgactcag tcaatcggag gactggcgcc
     6541 ccgagtgagg ggttgtgggc tcttttattg agctcgggga gcagaagcgc gcgaacagaa
     6601 gcgagaagcg aactgattgg ttagttcaaa taaggcacag ggtcatttca ggtccttggg
     6661 gcaccctgga aacatctgat ggttctctag aaactgctga gggctggacc gcatctgggg
     6721 accatctgtt cttggccctg agccggggca ggaactgctt accacagata tcctgtttgg
     6781 cccatattca gctgttccat ctgttcttgg ccctgagccg gggcaggaac tgcttaccac
     6841 agatatcctg tttggcccat attcagctgt tccatctgtt cctgaccttg atctgaactt
     6901 ctctattctc agttatgtat ttttccatgc cttgcaaaat ggcgttactt aagctagctt
     6961 gccaaaccta caggtggggt ctttcattcc cccctttttc tggagactaa ataaaatctt
     7021 ttattttatc gatgtttggc cgaggcggcc accggggatc tcaattaatc aacaggtccc
     7081 caaggtcgta ggagtcgaag agatcgctga tgccctcacc cgcctccaag ccccacaggt
     7141 agtcgtcctg gtccaaggat ggggagaagc tgatcagagg ggagctgcac gccagggtcg
     7201 gggacaggaa ctggtcctca gtctgctgca ggagtgggtg cggcagctcc agcaggctgt
     7261 cagtagcctc caaggggacc agggatggag gcggtggggc ctgctggggg gttggcgctg
     7321 gtgctggcac tgaggatgct gtgggctcca tgatgctagg gtcggtgctg ctgctgggct
     7381 gggcagagtc agggctgggg cagagggtgg aggtagaggg gagaggctcc tcggaaggac
     7441 tgtccggctc ctgcacctcc tctgggcaca ggtagacttc gatgggccct tgggtgctct
     7501 tgagatatat ctgcaggttg tcctcagtcc tgtcgggcac ttccagtctc gtctgcggag
     7561 gggccttgac ggcaatcact gtctgctcct taaagttgcc aacagcacgg atatcctggt
     7621 aagtcacata ggccagcctc ttgttggcct tgtcctcagt caggtgcttg aagctcagag
     7681 agcagctctg gatgagctgg tccaaggcct gctccgtgtt catcagctcc ttcagctcct
     7741 gccccagctg ttgctgcttc ccaggtctgg tggggtcttc aaacattccc ctgcctaccc
     7801 actggatgtt gttcttggcc ttcttgcgga tgagctggat gccttccagc acgttggtga
     7861 tgtcatagat gcgccgcttc tgcacgtcca gcacctcagc ggcccagttc aggtccagga
     7921 ccccatcctc tgactcgctc aggaggtaaa tgaacttctt ggtgagcagc cccagcgaag
     7981 tgtcataccg agtcttctcc ccgggggatt tgggggtttt ggggctgggg aggccatcca
     8041 ctctgatgca cttccccttg ggggttggga actcagggac gacgggcctc ccaatcccct
     8101 ccagatccag cttccttttg gccggcagcc ggcctgccgg caggcatcgc acaacttggc
     8161 cctcgggtcc gtggggagtg gcgtcgaggc aggtgcctgg cgccgctgcg ggaggcgccg
     8221 tctgcgggta cagcggtgtg tagtaggtag cagtagctgg gcagagctgg gggctgctga
     8281 ggccggatgg ccacagctct gtggggctca tcgcgggcac caccttcggg gtctgcccag
     8341 cggccgaagc caaggcccgg ggcccttgca gccgatccgc tcgagatctg agtccggact
     8401 tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggac
//
