LOCUS       sequence-396606-        5695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing human E2F3 protein with FLAG-tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5695)
  AUTHORS   Wang ME, Chen J, Lu Y, Bawcom AR, Wu J, Ou J, Asara JM, Armstrong 
            AJ, Wang Q, Li L, Wang Y, Huang J, Chen M
  TITLE     RB1-deficient prostate tumor growth and metastasis are vulnerable to
            ferroptosis induction via the E2F/ACSL4 axis.
  JOURNAL   J Clin Invest. 2023 Mar 16:e166647. doi: 10.1172/JCI166647.
  PUBMED    36928314
REFERENCE   2  (bases 1 to 5695)
  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: "J Clin 
            Invest. 2023 Mar 16:e166647. doi: 10.1172/JCI166647."
COMMENT     Sequence Label: pCMV-Tag2B-E2F3
FEATURES             Location/Qualifiers
     source          1..5695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        67..370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        371..574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     525..545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        620..638
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      673..682
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             682..705
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2124..2140)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2125..2141)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2196..2215)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2197..2215)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2489..2610
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2526..2545)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2580..2599
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2617..3072)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2754..2775)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2966..2985
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3099..3203
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3205..3562
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3205..3396
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3228..3248)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3413..3548
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3597..4391
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3597..4391
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3651..3670)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4261..4280
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4579..4598)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4623..4670
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4999..5587
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5488..5507
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcattagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
       61 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
      121 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      181 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      241 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      301 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      361 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      421 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      481 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      541 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      601 gcgattacgc caagctcgaa attaaccctc actaaaggga acaaaagctg gagctccacc
      661 gcggtggcgg ccgccaccat ggattacaag gatgacgacg ataagagccc gggcggatcc
      721 agaaagggaa tccagcccgc tctggagcag tacctggtga ccgccggggg tggggagggg
      781 gcggctgtcg tcgccgccgc cgctgcagcc tccatggaca aaagggcact gctagccagc
      841 cccggcttcg ccgccgccgc cgccgctgcc gccgccccgg gcgcgtacat ccagatcctc
      901 accacgaaca cttccaccac ctcctgttcc tcctccctcc aaagcggcgc cgtagccgcc
      961 ggccccctcc tccccagtgc ccccggcgcg gagcagaccg ccggcagcct cctctacacc
     1021 acgccgcacg gaccctccag cagagccggg ctgctgcagc agccaccagc gctgggacgc
     1081 ggcggcagcg gcggcggcgg cggccctccg gcaaagcgaa ggctggagct aggagaaagc
     1141 ggtcatcagt acctctcaga tggtttaaaa acccccaagg gcaaaggaag agctgcacta
     1201 cgaagtccag atagtccaaa aactccaaaa tctccctcag aaaaaacgcg gtatgatacg
     1261 tctcttggtc tgctcaccaa gaagttcatt cagctcctga gccagtcacc cgatggggta
     1321 ttggatttga acaaggcagc agaagtgcta aaagtgcaaa agagaaggat ttatgatatc
     1381 accaacgttc tggaaggcat ccacctcatt aagaagaagt ctaaaaacaa cgtccaatgg
     1441 atgggctgca gtctgtctga ggatgggggc atgctggccc agtgtcaagg cctgtcaaaa
     1501 gaagtgaccg agctcagtca ggaagagaag aaattagatg aactgatcca aagctgcacc
     1561 ctggacctca aactgttaac cgaggattca gagaatcaaa ggttagctta tgttacatat
     1621 caagatattc gaaaaattag tggccttaaa gaccaaactg ttatagttgt gaaagcccct
     1681 ccagaaacaa gacttgaagt gcctgactca atagagagcc tacaaataca tttggcaagt
     1741 acccaagggc ccattgaggt ttacttatgt ccagaagaga ctgaaacaca cagtccaatg
     1801 aaaacaaaca accaagacca caatgggaat atccctaaac ccgcttccaa agacttggct
     1861 tcaaccaact caggacatag cgattgctca gtttctatgg gaaacctttc tcctctggcc
     1921 tccccagcca acctcttaca gcagactgag gaccaaattc cttccaacct agaaggaccg
     1981 tttgtgaact tactgcctcc cctgctgcaa gaggactatc tcctgagcct cggggaggag
     2041 gaaggcatca gcgatctctt cgatgcttac gatttggaaa agctcccact ggtggaagac
     2101 ttcatgtgta gttgaaagct tatcgatacc gtcgacctcg agggggggcc cggtacctta
     2161 attaattaag gtaccaggta agtgtaccca attcgcccta tagtgagtcg tattacaatt
     2221 cactcgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggaga tccaattttt
     2281 aagtgtataa tgtgttaaac tactgattct aattgtttgt gtattttaga ttcacagtcc
     2341 caaggctcat ttcaggcccc tcagtcctca cagtctgttc atgatcataa tcagccatac
     2401 cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa
     2461 acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa
     2521 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     2581 tggtttgtcc aaactcatca atgtatctta acgcgtaaat tgtaagcgtt aatattttgt
     2641 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     2701 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     2761 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     2821 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     2881 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     2941 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     3001 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     3061 tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3121 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     3181 aataatattg aaaaaggaag aatcctgagg cggaaagaac cagctgtgga atgtgtgtca
     3241 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3301 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3361 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3421 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3481 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
     3541 tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga
     3601 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     3661 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     3721 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag
     3781 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     3841 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     3901 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     3961 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4021 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagaac
     4081 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg
     4141 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4201 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4261 cgttggctac ccgtgatatt gctgaagaac ttggcggcga atgggctgac cgcttcctcg
     4321 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4381 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4441 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4501 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4561 ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat
     4621 gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg
     4681 ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata
     4741 cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct
     4801 cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta
     4861 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     4921 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     4981 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     5041 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     5101 tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc
     5161 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     5221 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     5281 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     5341 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     5401 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     5461 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     5521 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     5581 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     5641 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcc
//
