LOCUS       sequence-396604-        5255 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing luciferase driven by human ACSL4 promoter (bases -277 to 
            -706 with respect to exon1 start site).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5255)
  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 5255)
  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/-277)
FEATURES             Location/Qualifiers
     source          1..5255
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     65..86
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    354..402
                     /note="synthetic polyadenylation signal"
     misc_feature    416..507
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     456..475
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     regulatory      1033..1042
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1039..2691
                     /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(1105..1123)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    2732..2853
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2743..2762)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2797..2816
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3101..3118)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3272..3860)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3352..3371)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4031..4822,4823..4891))
                     /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     4654..4673
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4892..4996)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      join(5023..5255,1..223)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5110..5129)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
       61 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
      121 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
      181 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttgccattc gccattcagg
      241 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagcccaag
      301 ctaccatgat aagtaagtaa tattaaggta cgggaggtac ttggagcggc cgcaataaaa
      361 tatctttatt ttcattacat ctgtgtgttg gttttttgtg tgaatcgata gtactaacat
      421 acgctctcca tcaaaacaaa acgaaacaaa acaaactagc aaaataggct gtccccagtg
      481 caagtgcagg tgccagaaca tttctctatc gataggtacc gagctcttac gcgtgctagc
      541 ccgggctcga gtaagaggca agataaacgg atgagaggca ttagttcagg cttacaggag
      601 caagatgctt ttagtcaaag tgttcaagat agctctctta aagaggagca gtgaaaccca
      661 aaaatgagtg tcttttgagg gcagctggaa gatgacccga ggggaacttt tatcctgggg
      721 tggagtcctg cgaagcaagc cgcaggtgag ggcgtgggcc aattctgcgc ctcactgctg
      781 ttaggcgcaa ggacatcccg aaacacactc aacgacacca gcagcgagac tgacagaagc
      841 ggatcgccag cgggagcaac aacttgaggc gagccgttag ccgcctcgcc ttcctcgcag
      901 cgcccctccc gcgccccggg aagcccgttc ggctccgcct tttcgactga cgtcccgggc
      961 gagcgggggc gggcgcgtgg gtgggccgag ccgtctgcag ccaaagcttg gcattccggt
     1021 actgttggta aagccaccat ggaagacgcc aaaaacataa agaaaggccc ggcgccattc
     1081 tatccgctgg aagatggaac cgctggagag caactgcata aggctatgaa gagatacgcc
     1141 ctggttcctg gaacaattgc ttttacagat gcacatatcg aggtggacat cacttacgct
     1201 gagtacttcg aaatgtccgt tcggttggca gaagctatga aacgatatgg gctgaataca
     1261 aatcacagaa tcgtcgtatg cagtgaaaac tctcttcaat tctttatgcc ggtgttgggc
     1321 gcgttattta tcggagttgc agttgcgccc gcgaacgaca tttataatga acgtgaattg
     1381 ctcaacagta tgggcatttc gcagcctacc gtggtgttcg tttccaaaaa ggggttgcaa
     1441 aaaattttga acgtgcaaaa aaagctccca atcatccaaa aaattattat catggattct
     1501 aaaacggatt accagggatt tcagtcgatg tacacgttcg tcacatctca tctacctccc
     1561 ggttttaatg aatacgattt tgtgccagag tccttcgata gggacaagac aattgcactg
     1621 atcatgaact cctctggatc tactggtctg cctaaaggtg tcgctctgcc tcatagaact
     1681 gcctgcgtga gattctcgca tgccagagat cctatttttg gcaatcaaat cattccggat
     1741 actgcgattt taagtgttgt tccattccat cacggttttg gaatgtttac tacactcgga
     1801 tatttgatat gtggatttcg agtcgtctta atgtatagat ttgaagaaga gctgtttctg
     1861 aggagccttc aggattacaa gattcaaagt gcgctgctgg tgccaaccct attctccttc
     1921 ttcgccaaaa gcactctgat tgacaaatac gatttatcta atttacacga aattgcttct
     1981 ggtggcgctc ccctctctaa ggaagtcggg gaagcggttg ccaagaggtt ccatctgcca
     2041 ggtatcaggc aaggatatgg gctcactgag actacatcag ctattctgat tacacccgag
     2101 ggggatgata aaccgggcgc ggtcggtaaa gttgttccat tttttgaagc gaaggttgtg
     2161 gatctggata ccgggaaaac gctgggcgtt aatcaaagag gcgaactgtg tgtgagaggt
     2221 cctatgatta tgtccggtta tgtaaacaat ccggaagcga ccaacgcctt gattgacaag
     2281 gatggatggc tacattctgg agacatagct tactgggacg aagacgaaca cttcttcatc
     2341 gttgaccgcc tgaagtctct gattaagtac aaaggctatc aggtggctcc cgctgaattg
     2401 gaatccatct tgctccaaca ccccaacatc ttcgacgcag gtgtcgcagg tcttcccgac
     2461 gatgacgccg gtgaacttcc cgccgccgtt gttgttttgg agcacggaaa gacgatgacg
     2521 gaaaaagaga tcgtggatta cgtcgccagt caagtaacaa ccgcgaaaaa gttgcgcgga
     2581 ggagttgtgt ttgtggacga agtaccgaaa ggtcttaccg gaaaactcga cgcaagaaaa
     2641 atcagagaga tcctcataaa ggccaagaag ggcggaaaga tcgccgtgta attctagagt
     2701 cggggcggcc ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc
     2761 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     2821 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     2881 tttcaggttc agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt
     2941 ggtaaaatcg ataaggatcc gtcgaccgat gcccttgaga gccttcaacc cagtcagctc
     3001 cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat
     3061 gcaactcgta ggacaggtgc cggcagcgct cttccgcttc ctcgctcact gactcgctgc
     3121 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     3181 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     3241 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     3301 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     3361 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     3421 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     3481 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     3541 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     3601 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     3661 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     3721 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     3781 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     3841 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     3901 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     3961 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     4021 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     4081 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     4141 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     4201 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     4261 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     4321 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     4381 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     4441 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     4501 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     4561 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     4621 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     4681 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     4741 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     4801 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     4861 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     4921 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     4981 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc tgtagcggcg
     5041 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     5101 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     5161 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5221 accccaaaaa acttgattag ggtgatggtt cacgt
//
