LOCUS       sequence-396602-        5509 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing luciferase driven by human ACSL4 promoter (bases -1 to 
            -706 with respect to exon1 start site).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5509)
  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 5509)
  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: pGL3-ACSL4-(-706)
FEATURES             Location/Qualifiers
     source          1..5509
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     61..82
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    350..398
                     /note="synthetic polyadenylation signal"
     misc_feature    412..503
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     452..471
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     regulatory      1283..1292
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1289..2941
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(1355..1373)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    2982..3103
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2993..3012)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3047..3066
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3351..3368)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3522..4110)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3602..3621)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4281..5072,5073..5141))
                     /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     4904..4923
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5142..5246)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(5273..5509,1..219)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5360..5379)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
       61 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      121 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      181 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttg ccattcgcca ttcaggctgc
      241 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag cccaagctac
      301 catgataagt aagtaatatt aaggtacggg aggtacttgg agcggccgca ataaaatatc
      361 tttattttca ttacatctgt gtgttggttt tttgtgtgaa tcgatagtac taacatacgc
      421 tctccatcaa aacaaaacga aacaaaacaa actagcaaaa taggctgtcc ccagtgcaag
      481 tgcaggtgcc agaacatttc tctatcgata ggtaccgagc tcttacgcgt gctagcccgg
      541 gctcgagtaa gaggcaagat aaacggatga gaggcattag ttcaggctta caggagcaag
      601 atgcttttag tcaaagtgtt caagatagct ctcttaaaga ggagcagtga aacccaaaaa
      661 tgagtgtctt ttgagggcag ctggaagatg acccgagggg aacttttatc ctggggtgga
      721 gtcctgcgaa gcaagccgca ggtgagggcg tgggccaatt ctgcgcctca ctgctgttag
      781 gcgcaaggac atcccgaaac acactcaacg acaccagcag cgagactgac agaagcggat
      841 cgccagcggg agcaacaact tgaggcgagc cgttagccgc ctcgccttcc tcgcagcgcc
      901 cctcccgcgc cccgggaagc ccgttcggct ccgccttttc gactgacgtc ccgggcgagc
      961 gggggcgggc gcgtgggtgg gccgagccgt ctgcagccag cgattcggct ggctctgcca
     1021 caccaccgcg cgcccccgct ccgcccgccc ctccgggcgc gtcttttccg ggctcgcgct
     1081 gagtcccgcc tccgccggct gtccgggtgc gcgcgcgccg ctgcggcttt ttctctggcc
     1141 tccgccgcgc gctcctcctc gtcccagcgc tagcgggcac gcggttcctt tttgcgagct
     1201 ttccgagtgc caggcgccgg ccggctgcga agacgcggtg ggccgcccct ccgaagcttg
     1261 gcattccggt actgttggta aagccaccat ggaagacgcc aaaaacataa agaaaggccc
     1321 ggcgccattc tatccgctgg aagatggaac cgctggagag caactgcata aggctatgaa
     1381 gagatacgcc ctggttcctg gaacaattgc ttttacagat gcacatatcg aggtggacat
     1441 cacttacgct gagtacttcg aaatgtccgt tcggttggca gaagctatga aacgatatgg
     1501 gctgaataca aatcacagaa tcgtcgtatg cagtgaaaac tctcttcaat tctttatgcc
     1561 ggtgttgggc gcgttattta tcggagttgc agttgcgccc gcgaacgaca tttataatga
     1621 acgtgaattg ctcaacagta tgggcatttc gcagcctacc gtggtgttcg tttccaaaaa
     1681 ggggttgcaa aaaattttga acgtgcaaaa aaagctccca atcatccaaa aaattattat
     1741 catggattct aaaacggatt accagggatt tcagtcgatg tacacgttcg tcacatctca
     1801 tctacctccc ggttttaatg aatacgattt tgtgccagag tccttcgata gggacaagac
     1861 aattgcactg atcatgaact cctctggatc tactggtctg cctaaaggtg tcgctctgcc
     1921 tcatagaact gcctgcgtga gattctcgca tgccagagat cctatttttg gcaatcaaat
     1981 cattccggat actgcgattt taagtgttgt tccattccat cacggttttg gaatgtttac
     2041 tacactcgga tatttgatat gtggatttcg agtcgtctta atgtatagat ttgaagaaga
     2101 gctgtttctg aggagccttc aggattacaa gattcaaagt gcgctgctgg tgccaaccct
     2161 attctccttc ttcgccaaaa gcactctgat tgacaaatac gatttatcta atttacacga
     2221 aattgcttct ggtggcgctc ccctctctaa ggaagtcggg gaagcggttg ccaagaggtt
     2281 ccatctgcca ggtatcaggc aaggatatgg gctcactgag actacatcag ctattctgat
     2341 tacacccgag ggggatgata aaccgggcgc ggtcggtaaa gttgttccat tttttgaagc
     2401 gaaggttgtg gatctggata ccgggaaaac gctgggcgtt aatcaaagag gcgaactgtg
     2461 tgtgagaggt cctatgatta tgtccggtta tgtaaacaat ccggaagcga ccaacgcctt
     2521 gattgacaag gatggatggc tacattctgg agacatagct tactgggacg aagacgaaca
     2581 cttcttcatc gttgaccgcc tgaagtctct gattaagtac aaaggctatc aggtggctcc
     2641 cgctgaattg gaatccatct tgctccaaca ccccaacatc ttcgacgcag gtgtcgcagg
     2701 tcttcccgac gatgacgccg gtgaacttcc cgccgccgtt gttgttttgg agcacggaaa
     2761 gacgatgacg gaaaaagaga tcgtggatta cgtcgccagt caagtaacaa ccgcgaaaaa
     2821 gttgcgcgga ggagttgtgt ttgtggacga agtaccgaaa ggtcttaccg gaaaactcga
     2881 cgcaagaaaa atcagagaga tcctcataaa ggccaagaag ggcggaaaga tcgccgtgta
     2941 attctagagt cggggcggcc ggccgcttcg agcagacatg ataagataca ttgatgagtt
     3001 tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc
     3061 tattgcttta tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat
     3121 tcattttatg tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct
     3181 ctacaaatgt ggtaaaatcg ataaggatcc gtcgaccgat gcccttgaga gccttcaacc
     3241 cagtcagctc cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct
     3301 tctttatcat gcaactcgta ggacaggtgc cggcagcgct cttccgcttc ctcgctcact
     3361 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     3421 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     3481 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     3541 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     3601 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     3661 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     3721 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     3781 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     3841 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     3901 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     3961 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     4021 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     4081 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     4141 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     4201 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     4261 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     4321 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     4381 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     4441 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     4501 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     4561 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     4621 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     4681 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     4741 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     4801 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     4861 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     4921 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     4981 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     5041 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     5101 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     5161 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     5221 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc
     5281 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     5341 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     5401 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     5461 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtg
//
