LOCUS       sequence-215000-        6673 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3'UTR reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6673)
  AUTHORS   Zhou Q, Anderson C, Zhang H, Li X, Inglis F, Jayagopal A, Wang S
  TITLE     Repression of choroidal neovascularization through actin 
            cytoskeleton pathways by microRNA-24.
  JOURNAL   Mol Ther. 2014 Feb;22(2):378-389. doi: 10.1038/mt.2013.243. Epub 
            2013 Oct 17.
  PUBMED    24297048
REFERENCE   2  (bases 1 to 6673)
  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: 
            "10.1038/mt.2013.243"; journalName: "Mol Ther"; date: "2014-02"; 
            volume: "22"; issue: "2"; pages: "378-389"
COMMENT     Sequence Label: pmiR-report-hEZH2
FEATURES             Location/Qualifiers
     source          1..6673
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..204
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     155..175
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             229..1878
                     /codon_start=1
                     /gene="luc"
                     /product="firefly luciferase"
                     /label=luciferase
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIVPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKSKL"
     primer_bind     complement(295..313)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(2237..2254)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2237..2253)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2246..2268)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2462..2481)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2643..2959
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2810..2945
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3056..3655
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3056..3075)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3552..3572
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    3885..3966
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3922..3941)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        4054..4158
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4159..4227,4228..5019)
                     /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(4377..4396)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5190..5778
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5679..5698
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5932..5949
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6066..6087
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6102..6107,6108..6125,6126..6132)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6140..6156
                     /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     6145..6167
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6164..6180
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6164..6180
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        6200..6218
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     6200..6217
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     6230..6249
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6230..6248
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        6370..6673
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
       61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      181 tgggaggtct atataagcag agctggttta gtgaaccgtg gatccaccat ggaagacgcc
      241 aaaaacataa agaaaggccc ggcgccattc tatcctctag aggatggaac cgctggagag
      301 caactgcata aggctatgaa gagatacgcc ctggttcctg gaacaattgc ttttacagat
      361 gcacatatcg aggtgaacat cacgtacgcg gaatacttcg aaatgtccgt tcggttggca
      421 gaagctatga aacgatatgg gctgaataca aatcacagaa tcgtcgtatg cagtgaaaac
      481 tctcttcaat tctttatgcc ggtgttgggc gcgttattta tcggagttgc agttgcgccc
      541 gcgaacgaca tttataatga acgtgaattg ctcaacagta tgaacatttc gcagcctacc
      601 gtagtgtttg tttccaaaaa ggggttgcaa aaaattttga acgtgcaaaa aaaattacca
      661 ataatccaga aaattattat catggattct aaaacggatt accagggatt tcagtcgatg
      721 tacacgttcg tcacatctca tctacctccc ggttttaatg aatacgattt tgtaccagag
      781 tcctttgatc gtgacaaaac aattgcactg ataatgaatt cctctggatc tactgggtta
      841 cctaagggtg tggcccttcc gcatagaact gcctgcgtca gattctcgca tgccagagat
      901 cctatttttg gcaatcaaat cgttccggat actgcgattt taagtgttgt cccattccat
      961 cacggttttg gaatgtttac tacactcgga tatttgatat gtggatttcg agtcgtctta
     1021 atgtatagat ttgaagaaga gctgttttta cgatcccttc aggattacaa aattcaaagt
     1081 gcgttgctag taccaaccct attttcattc ttcgccaaaa gcactctgat tgacaaatac
     1141 gatttatcta atttacacga aattgcttct gggggcgcac ctctttcgaa agaagtcggg
     1201 gaagcggttg caaaacgctt ccatcttcca gggatacgac aaggatatgg gctcactgag
     1261 actacatcag ctattctgat tacacccgag ggggatgata aaccgggcgc ggtcggtaaa
     1321 gttgttccat tttttgaagc gaaggttgtg gatctggata ccgggaaaac gctgggcgtt
     1381 aatcagagag gcgaattatg tgtcagagga cctatgatta tgtccggtta tgtaaacaat
     1441 ccggaagcga ccaacgcctt gattgacaag gatggatggc tacattctgg agacatagct
     1501 tactgggacg aagacgaaca cttcttcata gttgaccgct tgaagtcttt aattaaatac
     1561 aaaggatatc aggtggcccc cgctgaattg gaatcgatat tgttacaaca ccccaacatc
     1621 ttcgacgcgg gcgtggcagg tcttcccgac gatgacgccg gtgaacttcc cgccgccgtt
     1681 gttgttttgg agcacggaaa gacgatgacg gaaaaagaga tcgtggatta cgtcgccagt
     1741 caagtaacaa ccgcgaaaaa gttgcgcgga ggagttgtgt ttgtggacga agtaccgaaa
     1801 ggtcttaccg gaaaactcga cgcaagaaaa atcagagaga tcctcataaa ggccaagaag
     1861 ggcggaaagt ccaaattgct cgagtgatga aagctgcgca ctagtgaggg agctcttccc
     1921 cttctctctg aaacagctgc cttagcttca ggaaccttga gtactgtggg caatttagaa
     1981 aacggaaatg cagtttgaaa ttctgaattt gcaaagtact gtaacagtaa tttatagtaa
     2041 tgttttaaaa tcaacttttt attgccttct caccagctgc aaagtgtttt gtaccagtga
     2101 gtttttgcaa taatgcagta tggtacattt ttcaaatttg aataaagaat acttgaactt
     2161 gttgaagctt aataaaggat cttttatttt cattggatct gtgtgttggt tttttgtatg
     2221 cggccgctag cttggcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     2281 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     2341 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga
     2401 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatatgg tgcactctca
     2461 gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg
     2521 acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct
     2581 ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaaga
     2641 ttggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     2701 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     2761 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     2821 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     2881 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     2941 aggcctaggc ttttgcaaaa agctagcttg catgcctgca ggtcggccgc cacgaccggt
     3001 gccgccacca tcccctgacc cacgcccctg acccctcaca aggagacgac cttccatgac
     3061 cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtcccccggg ccgtacgcac
     3121 cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgacc cggaccgcca
     3181 catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg
     3241 caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt
     3301 cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg
     3361 gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc
     3421 gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc
     3481 cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct tcctggagac
     3541 ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca ccgccgacgt
     3601 cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg cctgacgccc
     3661 gccccacgac ccgcagcgcc cgaccgaaag gagcgcacga ccccatggct ccgaccgaag
     3721 ccacccgggg cggccccgcc gaccccgcac ccgcccccga ggcccaccga ctctagagga
     3781 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
     3841 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
     3901 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     3961 cactgcaatc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     4021 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     4081 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     4141 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     4201 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     4261 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     4321 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     4381 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     4441 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     4501 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     4561 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     4621 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     4681 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     4741 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     4801 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     4861 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     4921 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     4981 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     5041 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5101 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     5161 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     5221 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt ttttgccgga tcaagagcta
     5281 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     5341 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     5401 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     5461 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     5521 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5581 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5641 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5701 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5761 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5821 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     5881 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca
     5941 gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     6001 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     6061 gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt tatgcttccg
     6121 gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa cagctatgac
     6181 atgattacga attgcaacga tttaggtgac actatagaag agaaggaatt aatacgactc
     6241 actataggga gagagagaga attaccctca ctaaagggag gagaagcatg aattcaaggt
     6301 accagatctt agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat
     6361 ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc
     6421 ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
     6481 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
     6541 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
     6601 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
     6661 cgctattacc atg
//
