LOCUS       sequence-231676-        8177 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual luciferase assay for miRNA-targeted UTR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8177)
  AUTHORS   Kutsche LK, Gysi DM, Fallmann J, Lenk K, Petri R, Swiersy A, Klapper
            SD, Pircs K, Khattak S, Stadler PF, Jakobsson J, Nowick K, Busskamp 
            V
  TITLE     Combined Experimental and System-Level Analyses Reveal the Complex 
            Regulatory Network of miR-124 during Human Neurogenesis.
  JOURNAL   Cell Syst. 2018 Sep 28. pii: S2405-4712(18)30358-2. doi: 
            10.1016/j.cels.2018.08.011.
  PUBMED    30292704
REFERENCE   2  (bases 1 to 8177)
  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: "Cell Syst. 
            2018 Sep 28. pii: S2405-4712(18)30358-2. doi: 
            10.1016/j.cels.2018.08.011."
COMMENT     Sequence Label: pmiRGLO-ATF5-201
FEATURES             Location/Qualifiers
     source          1..8177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        45..544
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(80..103)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     477..496
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      585..594
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             591..2243
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     polyA_signal    3238..3359
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3249..3268)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3303..3322
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3604..3961
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3604..3795
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3627..3647)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3812..3947
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      4006..4015
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4012..4944
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             4996..5787
                     /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="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
                     LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
                     HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
                     LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
                     ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5047..5066)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5657..5676
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5851..5899
                     /note="synthetic polyadenylation signal"
     promoter        6055..6159
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6160..6228,6229..7020)
                     /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(6378..6397)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(7178..7766)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7258..7277)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    7882..8165
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
ORIGIN
        1 tactccggcc acgatgcgtc cggcgtagag gatcgagatc taccgggtag gggaggcgct
       61 tttcccaagg cagtctggag catgcgcttt agcagccccg ctgggcactt ggcgctacac
      121 aagtggcctc tggcctcgca cacattccac atccaccggt aggcgccaac cggctccgtt
      181 ctttggtggc cccttcgcgc caccttctac tcctccccta gtcaggaagt tcccccccgc
      241 cccgcagctc gcgtcgtgca ggacgtgaca aatggaagta gcacgtctca ctagtctcgt
      301 gcagatggac agcaccgctg agcaatggaa gcgggtaggc ctttggggca gcggccaata
      361 gcagctttgc tccttcgctt tctgggctca gaggctggga aggggtgggt ccgggggcgg
      421 gctcaggggc gggctcaggg gcggggcggg cgcccgaagg tcctccggag gcccggcatt
      481 ctgcacgctt caaaagcgca cgtctgccgc gctgttctcc tcttcctcat ctccgggcct
      541 ttcgacctgc agcccaagct tggcaatccg gtactgttgg taaagccacc atggaagatg
      601 ccaaaaacat taagaagggc ccagcgccat tctacccact cgaagacggg accgccggcg
      661 agcagctgca caaagccatg aagcgctacg ccctggtgcc cggcaccatc gcctttaccg
      721 acgcacatat cgaggtggac attacctacg ccgagtactt cgagatgagc gttcggctgg
      781 cagaagctat gaagcgctat gggctgaata caaaccatcg gatcgtggtg tgcagcgaga
      841 atagcttgca gttcttcatg cccgtgttgg gtgccctgtt catcggtgtg gctgtggccc
      901 cagctaacga catctacaac gagcgcgagc tgctgaacag catgggcatc agccagccca
      961 ccgtcgtatt cgtgagcaag aaagggctgc aaaagatcct caacgtgcaa aagaagctac
     1021 cgatcataca aaagatcatc atcatggata gcaagaccga ctaccagggc ttccaaagca
     1081 tgtacacctt cgtgacttcc catttgccac ccggcttcaa cgagtacgac ttcgtgcccg
     1141 agagcttcga ccgggacaaa accatcgccc tgatcatgaa cagtagtggc agtaccggat
     1201 tgcccaaggg cgtagcccta ccgcaccgca ccgcttgtgt ccgattcagt catgcccgcg
     1261 accccatctt cggcaaccag atcatccccg acaccgctat cctcagcgtg gtgccatttc
     1321 accacggctt cggcatgttc accacgctgg gctacttgat ctgcggcttt cgggtcgtgc
     1381 tcatgtaccg cttcgaggag gagctattct tgcgcagctt gcaagactat aagattcaat
     1441 ctgccctgct ggtgcccaca ctatttagct tcttcgctaa gagcactctc atcgacaagt
     1501 acgacctaag caacttgcac gagatcgcca gcggcggggc gccgctcagc aaggaggtag
     1561 gtgaggccgt ggccaaacgc ttccacctac caggcatccg ccagggctac ggcctgacag
     1621 aaacaaccag cgccattctg atcacccccg aaggggacga caagcctggc gcagtaggca
     1681 aggtggtgcc cttcttcgag gctaaggtgg tggacttgga caccggtaag acactgggtg
     1741 tgaaccagcg cggcgagctg tgcgtccgtg gccccatgat catgagcggc tacgttaaca
     1801 accccgaggc tacaaacgct ctcatcgaca aggacggctg gctgcacagc ggcgacatcg
     1861 cctactggga cgaggacgag cacttcttca tcgtggaccg gctgaagagc ctgatcaaat
     1921 acaagggcta ccaggtagcc ccagccgaac tggagagcat cctgctgcaa caccccaaca
     1981 tcttcgacgc cggggtcgcc ggcctgcccg acgacgatgc cggcgagctg cccgccgcag
     2041 tcgtcgtgct ggaacacggt aaaaccatga ccgagaagga gatcgtggac tatgtggcca
     2101 gccaggttac aaccgccaag aagctgcgcg gtggtgttgt gttcgtggac gaggtgccta
     2161 aaggactgac cggcaagttg gacgcccgca agatccgcga gattctcatt aaggccaaga
     2221 agggcggcaa gatcgccgtg taattctagt tgtttaaacg agctcgaagg gcaggggtgt
     2281 ggcttctggg ggctggtctt cagctctggc gccttcatcc ccctgcctct accttcattc
     2341 caaacccctc tcggccgggt gcagtggctt atgcttgtaa tcccagcact ttgggaggcc
     2401 aaggcaggag gatcgtttga ggccaggagg tcaataccag cctgggcaac atagtaagac
     2461 cctgtctcta ttaaaaaaaa aaaatcaacc cttcttcccc accaaaccac ccaactcctc
     2521 tctactctta tccttttatc ctctgtctct gcttatcacc tctcttgcgt atttctggat
     2581 ctccttccct cctttctcgt ccaaatcatg aaatgtttgg ccttagtcaa tgtctatgcc
     2641 cgtcacataa cagccgaggc accgaggccc acagggaagc agctgggagc ttggaaacct
     2701 ggtctcttga atttcaaacc tggtttctta caggtggttg tctggggtgg gtggagtggc
     2761 gacaggatag agctgaagga ctatgcaaat gaggaagtaa gtcagggcgg gctttgagaa
     2821 ggggacccat atcctacagg caaaaagcag gctaggtgac cttgggacac tacgctaagg
     2881 gagggaggct aaaggcggcc aggtttgcag tgcgggaaga tgagcaggcc agtgggagga
     2941 ggggcagggc agggctgtag ttggtgactg ggtgttcatt ttagctctaa gaaaaaaaat
     3001 cagtgtttcg tgaaggtgtt ggagaggggc tgtgtctggg tgagggatgg cggggtactg
     3061 atttttttgg gaggttatga gcaaaaataa aacgaaacat ttcctctgga gtcgacctgc
     3121 aggcatgcaa gctgatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc
     3181 caccgctgag caataactag cataacccct tggggcggcc gcttcgagca gacatgataa
     3241 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     3301 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     3361 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt
     3421 taagcaagta aaacctctac aaatgtggta aaatcgaatt ttaacaaaat attaacgctt
     3481 acaatttcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     3541 gcggatctgc gcagcaccat ggcctgaaat aacctctgaa agaggaactt ggttaggtac
     3601 cttctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     3661 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     3721 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3781 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     3841 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     3901 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     3961 aaagcttgat tcttctgaca caacagtctc gaaccaaagg ctggagccac catggcttcc
     4021 aaggtgtacg accccgagca acgcaaacgc atgatcactg ggcctcagtg gtgggctcgc
     4081 tgcaagcaaa tgaacgtgct ggactccttc atcaactact atgattccga gaagcacgcc
     4141 gagaacgccg tgatttttct gcatggtaac gctgcctcca gctacctgtg gaggcacgtc
     4201 gtgcctcaca tcgagcccgt ggctagatgc atcatccctg atctgatcgg aatgggtaag
     4261 tccggcaaga gcgggaatgg ctcatatcgc ctcctggatc actacaagta cctcaccgct
     4321 tggttcgagc tgctgaacct tccaaagaaa atcatctttg tgggccacga ctggggggct
     4381 tgtctggcct ttcactactc ctacgagcac caagacaaga tcaaggccat cgtccatgct
     4441 gagagtgtcg tggacgtgat cgagtcctgg gacgagtggc ctgacatcga ggaggatatc
     4501 gccctgatca agagcgaaga gggcgagaaa atggtgcttg agaataactt cttcgtcgag
     4561 accatgctcc caagcaagat catgcggaaa ctggagcctg aggagttcgc tgcctacctg
     4621 gagccattca aggagaaggg cgaggttaga cggcctaccc tctcctggcc tcgcgagatc
     4681 cctctcgtta agggaggcaa gcccgacgtc gtccagattg tccgcaacta caacgcctac
     4741 cttcgggcca gcgacgatct gcctaagatg ttcatcgagt ccgaccctgg gttcttttcc
     4801 aacgctattg tcgagggagc taagaagttc cctaacaccg agttcgtgaa ggtgaagggc
     4861 ctccacttca gccaggagga cgctccagat gaaatgggta agtacatcaa gagcttcgtg
     4921 gagcgcgtgc tgaagaacga gcagaccggt ggtgggagcg gaggtggcgg atcaggtggc
     4981 ggaggctccg gagggattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     5041 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     5101 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     5161 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     5221 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     5281 gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg
     5341 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     5401 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     5461 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     5521 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     5581 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     5641 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     5701 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     5761 tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag
     5821 cgacgcccaa cctgccatca cgatggccgc aataaaatat ctttattttc attacatctg
     5881 tgtgttggtt ttttgtgtga atcgatagcg ataaggatcc tctttgcgct tgcgttttcc
     5941 cttgtccaga tagcccagta gctgacattc atccggggtc agcaccgttt ctgcggactg
     6001 gctttctacg taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     6061 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     6121 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     6181 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     6241 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     6301 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     6361 agcactttca aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     6421 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     6481 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     6541 agtgataaca ctgcggccaa cttacttctg acaactatcg gaggaccgaa ggagctaacc
     6601 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     6661 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     6721 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     6781 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     6841 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     6901 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     6961 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     7021 ttcgaaatga ccgaccaagc gacgcccaac cggtatcagc tcactcaaag gcggtaatac
     7081 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     7141 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     7201 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     7261 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     7321 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     7381 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     7441 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     7501 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     7561 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga
     7621 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     7681 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     7741 ttacgcgcag aaaaaaagga tttcaagaag atcctttgat cttttctacg gggtctgacg
     7801 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     7861 tcacctagat ccttttatag tccggaaata caggaacgca cgctggatgg cccttcgctg
     7921 ggatggtgaa accatgaaaa atggcagctt cagtggatta agtgggggta atgtggcctg
     7981 taccctctgg ttgcataggt attcatacgg ttaaaattta tcaggcgcga ttgcggcagt
     8041 ttttcgggtg gtttgttgcc atttttacct gtctgctgcc gtgatcgcgc tgaacgcgtt
     8101 ttagcggtgc gtacaattaa gggattatgg taaatccact tactgtctgc cctcgtagcc
     8161 atcgagataa accgcag
//
