LOCUS       sequence-473630-        6502 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transient overexpression of E2F3B in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6502)
  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 6502)
  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: pcDNA3 HA E2F3B
FEATURES             Location/Qualifiers
     source          1..6502
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      320..908
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     809..828
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1062..1079
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1196..1217
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1232..1237,1238..1255,1256..1262)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1270..1286
                     /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     1275..1297
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1294..1310
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1294..1310
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    1359..1480
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1370..1389)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1424..1443
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1654..2448)
                     /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(1765..1784)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2375..2394
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2515..2844)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2529..2664
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(2583..2602)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2829..2849
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2858..3286)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2968..2989)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3180..3199
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    3332..3556
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3545..3562
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     promoter        3582..3600
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3582..3599
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             complement(4719..4745)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             complement(4725..4751)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             complement(4725..4751)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(4773..4792)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4774..4792)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(4837..5040)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(4866..4886)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5041..5420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     5592..5611
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        5686..5790
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5791..5859,5860..6502,1..149)
                     /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(6009..6028)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
       61 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
      121 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
      181 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
      241 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
      301 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
      361 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
      421 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
      481 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
      541 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
      601 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
      661 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
      721 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
      781 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
      841 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
      901 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
      961 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     1021 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     1081 aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     1141 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     1201 tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt
     1261 tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg
     1321 ccaagctcta gctagaggtc gacggtatac agacatgata agatacattg atgagtttgg
     1381 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     1441 tgctttattt gtaaccatta taagctgcaa taaacaagtt ggggtgggcg aagaactcca
     1501 gcatgagatc cccgcgctgg aggatcatcc agccggcgtc ccggaaaacg attccgaagc
     1561 ccaacctttc atagaaggcg gcggtggaat cgaaatctcg tgatggcagg ttgggcgtcg
     1621 cttggtcggt catttcgaac cccagagtcc cgctcagaag aactcgtcaa gaaggcgata
     1681 gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa agcacgagga agcggtcagc
     1741 ccattcgccg ccaagctctt cagcaatatc acgggtagcc aacgctatgt cctgatagcg
     1801 gtccgccaca cccagccggc cacagtcgat gaatccagaa aagcggccat tttccaccat
     1861 gatattcggc aagcaggcat cgccatgggt cacgacgaga tcctcgccgt cgggcatgcg
     1921 cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc tgatgctctt cgtccagatc
     1981 atcctgatcg acaagaccgg cttccatccg agtacgtgct cgctcgatgc gatgtttcgc
     2041 ttggtggtcg aatgggcagg tagccggatc aagcgtatgc agccgccgca ttgcatcagc
     2101 catgatggat actttctcgg caggagcaag gtgagatgac aggagatcct gccccggcac
     2161 ttcgcccaat agcagccagt cccttcccgc ttcagtgaca acgtcgagca cagctgcgca
     2221 aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc tcgtcctgca gttcattcag
     2281 ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc ccctgcgctg acagccggaa
     2341 cacggcggca tcagagcagc cgattgtctg ttgtgcccag tcatagccga atagcctctc
     2401 cacccaagcg gccggagaac ctgcgtgcaa tccatcttgt tcaatcatgc gaaacgatcc
     2461 tcatcctgtc tcttgatcag atccgaaaat ggatatacaa gctcccggga gctttttgca
     2521 aaagcctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag aggcagaggc
     2581 ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga
     2641 actgggcgga gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta
     2701 attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct
     2761 ggttgctgac taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga
     2821 ctttccacac cctaactgac acacattcca cagaattaat tcgcgttaaa tttttgttaa
     2881 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
     2941 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
     3001 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
     3061 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
     3121 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
     3181 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
     3241 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtgggg atacccccta
     3301 gagccccagc tggttctttc cgcctcagaa gccatagagc ccaccgcatc cccagcatgc
     3361 ctgctattgt cttcccaatc ctcccccttg ctgtcctgcc ccaccccacc ccccagaata
     3421 gaatgacacc tactcagaca atgcgatgca atttcctcat tttattagga aaggacagtg
     3481 ggagtggcac cttccagggt caaggaaggc acgggggagg ggcaaacaac agatggctgg
     3541 caactagaag gcacagtcga ggctgatcag cgagctctag catttaggtg acactataga
     3601 atagggccct ctagatgcat gctcgagcgg ccgccagtgt gatggatatc tgcagaattt
     3661 caactacaca tgaagtcttc caccagtggg agcttttcca aatcgtaagc atcgaagaga
     3721 tcgctgatgc cttcctcctc cccgaggctc aggagatagt cctcttgcag caggggaggc
     3781 agtaagttca caaacggtcc ttctaggttg gaaggaattt ggtcctcagt ctgctgtaag
     3841 aggttggctg gggaggccag aggagaaagg tttcccatag aaactgagca atcgctatgt
     3901 cctgagttgg ttgaagccaa gtctttggaa gcgggtttag ggatattccc attgtggtct
     3961 tggttgtttg ttttcattgg actgtgtgtt tcagtctctt ctggacataa gtaaacctca
     4021 atgggccctt gggtacttgc caaatgtatt tgtaggctct ctattgagtc aggcacttca
     4081 agtcttgttt ctggaggggc tttcacaact ataacagttt ggtctttaag gccactaatt
     4141 tttcgaatat cttgatatgt aacataagct aacctttgat tctctgaatc ctcggttaac
     4201 agtttgaggt ccagggtgca gctttggatc agttcatcta atttcttctc ttcctgactg
     4261 agctcggtca cttcttttga caggccttga cactgggcca gcatgccccc atcctcagac
     4321 agactgcagc ccatccattg gacgttgttt ttagacttct tcttaatgag gtggatgcct
     4381 tccagaacgt tggtgatatc ataaatcctt ctcttttgca cttttagcac ttctgctgcc
     4441 ttgttcaaat ccaatacccc atcgggtgac tggctcagga gctgaatgaa cttcttggtg
     4501 agcagaccaa gagacgtatc ataccgcgtt ttttctgagg gagattttgg agtttttgga
     4561 ctatctggac ttcgtagtgc agctcttcct ttgcccttgg gggtttttaa accatctgag
     4621 aggtactgat gaccgctttc tcctagctcc agccttcgct ttgcctgctg ctgtaagggc
     4681 agaattccag cacactggcg gccgttacta gtggatccga gggaggcgta gtcgggcacg
     4741 tcgtaggggt aggccatggt aagcttgggt ctccctatag tgagtcgtat taatttcgat
     4801 aagccagtaa gcagtgggtt ctctagttag ccagagagct ctgcttatat agacctccca
     4861 ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac attttggaaa
     4921 gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt ggagacttgg
     4981 aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa accgcatcac
     5041 catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt cccataaggt
     5101 catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa tagggggcgt
     5161 acttggcata tgatacactt gatgtactgc caagtgggca gtttaccgta aatactccac
     5221 ccattgacgt caatggaaag tccctattgg cgttactatg ggaacatacg tcattattga
     5281 cgtcaatggg cgggggtcgt tgggcggtca gccaggcggg ccatttaccg taagttatgt
     5341 aacgcggaac tccatatatg ggctatgaac taatgacccc gtaattgatt actattaata
     5401 actagtcaat aatcaatgtc aacgcgtata tctggcccgt acatcgcgaa gcagcgcaaa
     5461 acgcctaacc ctaagcagat tcttcatgca attgtcggtc aagccttgcc ttgttgtagc
     5521 ttaaattttg ctcgcgcact actcagcgac ctccaacaca caagcaggga gcagatactg
     5581 gcttaactat gcggcatcag agcagattgt actgagagtg caccataggg gatcgggaga
     5641 tctcccgatc cgtcgacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     5701 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     5761 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     5821 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     5881 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     5941 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     6001 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     6061 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     6121 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     6181 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     6241 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     6301 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     6361 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     6421 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     6481 gataaatctg gagccggtga gc
//
