LOCUS       sequence-403634-        7642 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing luciferase driven by human ACSL4 promoter (bases -1 to 
            -2844 with respect to exon1 start site).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7642)
  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 7642)
  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-FL
FEATURES             Location/Qualifiers
     source          1..7642
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     200..221
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    489..537
                     /note="synthetic polyadenylation signal"
     misc_feature    551..642
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     591..610
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     regulatory      3555..3564
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3561..5213
                     /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(3627..3645)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    5254..5375
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5265..5284)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5319..5338
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5623..5640)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5794..6382)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5874..5893)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6553..7344,7345..7413))
                     /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     7176..7195
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7414..7518)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(7545..7642,1..358)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(join(7632..7642,1..9))
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
       61 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
      121 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
      181 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
      241 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
      301 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttgc
      361 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
      421 ttacgccagc ccaagctacc atgataagta agtaatatta aggtacggga ggtacttgga
      481 gcggccgcaa taaaatatct ttattttcat tacatctgtg tgttggtttt ttgtgtgaat
      541 cgatagtact aacatacgct ctccatcaaa acaaaacgaa acaaaacaaa ctagcaaaat
      601 aggctgtccc cagtgcaagt gcaggtgcca gaacatttct ctatcgatag gtaccgagct
      661 cttacgcgtg ctagcccggg ctcgagcccg tgaagtttaa cacgcaaagg cttgttacac
      721 agtcaatgaa gaacagttgt tttctgccct tccataaata actaatgcca agagcagcct
      781 ggctatctgt ttcagcttct tgttacagaa taataaaata ggcaaggcct aaatatcagt
      841 gcctgttaac agagctgtcc tttaaattct gtttagcttc taacagacta attctcatgc
      901 ttttagtact tattacattg cattttgaat ctgtcaattg cattgtacat tttcctaact
      961 cttccatttc catacgttta tctactccat ttctaattgg gttggtagtt ctgggtagga
     1021 ctcattacag aggcagctta atgtagcggc aagcaccctg gtctaggagt cataagcagt
     1081 gtagtaggaa attcaggctc tgttactact tagttgtgtg actggacaag acacttaacc
     1141 tgcatctatt tcttcatctg tgaaataggg ataataatac cctatcttcc taactaataa
     1201 acaacgttgc aaatgttctc tttaaaatgt cagacagctg cttgggaaat attttgctcc
     1261 tgggcagtaa tttctggaag gtttctgatg acaagaaatt aagcaaggtt tgttttgact
     1321 ccgaaaaagg aaagctggaa atagcgtggt tatacttggt ttcctttttg gaggtatagt
     1381 tcttctttcc tcttcaactt gggtgcagtt aaatgtggaa gaaggtgaga tgtggtaggt
     1441 gatgaagaca gccagaactg tacacactaa tgaatagctt aagtaataag agatcttgaa
     1501 aatccaagta gtttcttggt ctgataaaca tatctatatg tatatcatac acacatttgt
     1561 ataccaaaaa acggatagtt tcaggctata gtgatttctt tcctttatat cttgtagagc
     1621 tggaggaact tcagtaagct gaagactctg ttcagaggaa attacctaat tgaaaatgtc
     1681 tgggctaaga gaccgacaat aaaggaaact ggcaaaagta ctctttaatt acaagttctt
     1741 tttcatcttc atccagcaca ctgatttatg tgctacactg ggtttgtatt atatctatat
     1801 tttaattcca caactagaca ctaactttat tgagaacagc gatattcatt tcttcaattt
     1861 catttgtgtt aacccaactg taggtgatag tgctctgtac atcatagtta cccagtgaat
     1921 tacgtacctg acatatgatt gcttatttct ggtattaaaa caagatttca taaaataaac
     1981 atgacaatta aactcgttgt ttgaagtact gaactttgta ggtcaatttt gaaaaaggaa
     2041 gctacatgac tgtcttagca atatttatag gtatattata tgtaagtggg tgggtttctt
     2101 ggaaattgaa ttggtaatat ggctttagag tagtaacaca aaaactatag ctctgtgctc
     2161 tcagctaacc acttttagtg tcacctgggc tgcttattaa aaattcaaat ttcttatccc
     2221 tacccagatc agaatcaaaa tacctggggg ttgtagctcc aggtacctac atttcaacaa
     2281 gcaccccaga taattgtgat gcacactgaa gaactactct agcttgtaat ctcaggtggt
     2341 aaggcatttt atatatacat atatatatac acacacacaa gcacaaatgg taattctatg
     2401 ggaaaatgga aatcagcaag tgaagtgaaa tgtcaattac catcaggaac aggaagctag
     2461 ttcaccatta cgcatcaggg aactggaaga attcttgaga tttatttttt tgggtgatat
     2521 attcttggca gtttatcagg cctcatttgc aattatggtc tgtccctttt gaagccctca
     2581 gatagttcgc tggcatttta attttaatag caatttcttg catctgttgg aacagcattc
     2641 gtatgaccgg cttataagca atgatcaggg ctagtaagat gtcacaaagc tgcggtgact
     2701 tttccttatc agttgggggt ttttttgcca tgataccatc gtggaaatga aagacaacgt
     2761 gaaaatcctg cgtccttgtg atatttcatc agtgggagtg agcggggatc ttaacgcagt
     2821 aagaggcaag ataaacggat gagaggcatt agttcaggct tacaggagca agatgctttt
     2881 agtcaaagtg ttcaagatag ctctcttaaa gaggagcagt gaaacccaaa aatgagtgtc
     2941 ttttgagggc agctggaaga tgacccgagg ggaactttta tcctggggtg gagtcctgcg
     3001 aagcaagccg caggtgaggg cgtgggccaa ttctgcgcct cactgctgtt aggcgcaagg
     3061 acatcccgaa acacactcaa cgacaccagc agcgagactg acagaagcgg atcgccagcg
     3121 ggagcaacaa cttgaggcga gccgttagcc gcctcgcctt cctcgcagcg cccctcccgc
     3181 gccccgggaa gcccgttcgg ctccgccttt tcgactgacg tcccgggcga gcgggggcgg
     3241 gcgcgtgggt gggccgagcc gtctgcagcc agcgattcgg ctggctctgc cacaccaccg
     3301 cgcgcccccg ctccgcccgc ccctccgggc gcgtcttttc cgggctcgcg ctgagtcccg
     3361 cctccgccgg ctgtccgggt gcgcgcgcgc cgctgcggct ttttctctgg cctccgccgc
     3421 gcgctcctcc tcgtcccagc gctagcgggc acgcggttcc tttttgcgag ctttccgagt
     3481 gccaggcgcc ggccggctgc gaagacgcgg tgggccgccc ctccgaagct tggcattccg
     3541 gtactgttgg taaagccacc atggaagacg ccaaaaacat aaagaaaggc ccggcgccat
     3601 tctatccgct ggaagatgga accgctggag agcaactgca taaggctatg aagagatacg
     3661 ccctggttcc tggaacaatt gcttttacag atgcacatat cgaggtggac atcacttacg
     3721 ctgagtactt cgaaatgtcc gttcggttgg cagaagctat gaaacgatat gggctgaata
     3781 caaatcacag aatcgtcgta tgcagtgaaa actctcttca attctttatg ccggtgttgg
     3841 gcgcgttatt tatcggagtt gcagttgcgc ccgcgaacga catttataat gaacgtgaat
     3901 tgctcaacag tatgggcatt tcgcagccta ccgtggtgtt cgtttccaaa aaggggttgc
     3961 aaaaaatttt gaacgtgcaa aaaaagctcc caatcatcca aaaaattatt atcatggatt
     4021 ctaaaacgga ttaccaggga tttcagtcga tgtacacgtt cgtcacatct catctacctc
     4081 ccggttttaa tgaatacgat tttgtgccag agtccttcga tagggacaag acaattgcac
     4141 tgatcatgaa ctcctctgga tctactggtc tgcctaaagg tgtcgctctg cctcatagaa
     4201 ctgcctgcgt gagattctcg catgccagag atcctatttt tggcaatcaa atcattccgg
     4261 atactgcgat tttaagtgtt gttccattcc atcacggttt tggaatgttt actacactcg
     4321 gatatttgat atgtggattt cgagtcgtct taatgtatag atttgaagaa gagctgtttc
     4381 tgaggagcct tcaggattac aagattcaaa gtgcgctgct ggtgccaacc ctattctcct
     4441 tcttcgccaa aagcactctg attgacaaat acgatttatc taatttacac gaaattgctt
     4501 ctggtggcgc tcccctctct aaggaagtcg gggaagcggt tgccaagagg ttccatctgc
     4561 caggtatcag gcaaggatat gggctcactg agactacatc agctattctg attacacccg
     4621 agggggatga taaaccgggc gcggtcggta aagttgttcc attttttgaa gcgaaggttg
     4681 tggatctgga taccgggaaa acgctgggcg ttaatcaaag aggcgaactg tgtgtgagag
     4741 gtcctatgat tatgtccggt tatgtaaaca atccggaagc gaccaacgcc ttgattgaca
     4801 aggatggatg gctacattct ggagacatag cttactggga cgaagacgaa cacttcttca
     4861 tcgttgaccg cctgaagtct ctgattaagt acaaaggcta tcaggtggct cccgctgaat
     4921 tggaatccat cttgctccaa caccccaaca tcttcgacgc aggtgtcgca ggtcttcccg
     4981 acgatgacgc cggtgaactt cccgccgccg ttgttgtttt ggagcacgga aagacgatga
     5041 cggaaaaaga gatcgtggat tacgtcgcca gtcaagtaac aaccgcgaaa aagttgcgcg
     5101 gaggagttgt gtttgtggac gaagtaccga aaggtcttac cggaaaactc gacgcaagaa
     5161 aaatcagaga gatcctcata aaggccaaga agggcggaaa gatcgccgtg taattctaga
     5221 gtcggggcgg ccggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
     5281 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     5341 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     5401 tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac ctctacaaat
     5461 gtggtaaaat cgataaggat ccgtcgaccg atgcccttga gagccttcaa cccagtcagc
     5521 tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt cttctttatc
     5581 atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca ctgactcgct
     5641 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5701 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5761 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5821 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5881 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5941 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     6001 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     6061 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     6121 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6181 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6241 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6301 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6361 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6421 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6481 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6541 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6601 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6661 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6721 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6781 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6841 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6901 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6961 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     7021 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     7081 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     7141 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7201 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7261 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7321 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7381 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7441 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7501 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg
     7561 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     7621 cctagcgccc gctcctttcg ct
//
