LOCUS       sequence-269426-        6835 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3 repeats of BoxB and MS2 hairpins inserted into the 3'UTR of 
            Renilla luciferase in the psiCHECK3 plasmid for the Tethering assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6835)
  AUTHORS   Fischer JW, Busa VF, Shao Y, Leung AKL
  TITLE     Structure-Mediated RNA Decay by UPF1 and G3BP1.
  JOURNAL   Mol Cell. 2020 Apr 2;78(1):70-84.e6. doi: 
            10.1016/j.molcel.2020.01.021. Epub 2020 Feb 3.
  PUBMED    32017897
REFERENCE   2  (bases 1 to 6835)
  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: 
            "10.1016/j.molcel.2020.01.021"; journalName: "Mol Cell"; date: 
            "2020-04-2- 2"; volume: "78"; issue: "1"; pages: "70-84.e6"
COMMENT     Sequence Label: psi-CHECK3-3xBoxB-MS2
FEATURES             Location/Qualifiers
     source          1..6835
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        66..423
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        66..257
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(89..109)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      274..409
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          493..625
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        670..688
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      692..701
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             698..1633
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     misc_RNA        1687..1705
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        1795..1813
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        1903..1921
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     polyA_signal    2029..2077
                     /note="synthetic polyadenylation signal"
     promoter        2086..2837
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          2907..3039
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        3084..3102
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      3111..3120
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3117..4769
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /db_xref="GI:42718124"
                     /protein_id="AAS38485.1"
                     /translation="MADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             4076..4084
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    4794..4915
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4805..4824)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4859..4878
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5048..5152
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5153..5221,5222..6013)
                     /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(5371..5390)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6184..6772
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6673..6692
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgacagatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt
       61 accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
      121 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
      181 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
      241 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
      301 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
      361 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
      421 aaaaagcttg attcttctga cacaacagtc tcgaacttaa gctgcagaag ttggtcgtga
      481 ggcactgggc aggtaagtat caaggttaca agacaggttt aaggagacca atagaaactg
      541 ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg tcttactgac
      601 atccactttg cctttctctc cacaggtgtc cactcccagt tcaattacag ctcttaaggc
      661 tagagtactt aatacgactc actataggct agccaccatg gcttccaagg tgtacgaccc
      721 cgagcaacgc aaacgcatga tcactgggcc tcagtggtgg gctcgctgca agcaaatgaa
      781 cgtgctggac tccttcatca actactatga ttccgagaag cacgccgaga acgccgtgat
      841 ttttctgcat ggtaacgctg cctccagcta cctgtggagg cacgtcgtgc ctcacatcga
      901 gcccgtggct agatgcatca tccctgatct gatcggaatg ggtaagtccg gcaagagcgg
      961 gaatggctca tatcgcctcc tggatcacta caagtacctc accgcttggt tcgagctgct
     1021 gaaccttcca aagaaaatca tctttgtggg ccacgactgg ggggcttgtc tggcctttca
     1081 ctactcctac gagcaccaag acaagatcaa ggccatcgtc catgctgaga gtgtcgtgga
     1141 cgtgatcgag tcctgggacg agtggcctga catcgaggag gatatcgccc tgatcaagag
     1201 cgaagagggc gagaaaatgg tgcttgagaa taacttcttc gtcgagacca tgctcccaag
     1261 caagatcatg cggaaactgg agcctgagga gttcgctgcc tacctggagc cattcaagga
     1321 gaagggcgag gttagacggc ctaccctctc ctggcctcgc gagatccctc tcgttaaggg
     1381 aggcaagccc gacgtcgtcc agattgtccg caactacaac gcctaccttc gggccagcga
     1441 cgatctgcct aagatgttca tcgagtccga ccctgggttc ttttccaacg ctattgtcga
     1501 gggagctaag aagttcccta acaccgagtt cgtgaaggtg aagggcctcc acttcagcca
     1561 ggaggacgct ccagatgaaa tgggtaagta catcaagagc ttcgtggagc gcgtgctgaa
     1621 gaacgagcag taattctagg cgatcgctcg agccgtgtaa taattctagc caaaggttcc
     1681 tttgtcacat gaggatcacc catgtgaatt ggtggctggt gtggctaatg ccctatggcc
     1741 ctgaaaaagg gccactggag gatattcatg catctagcca aaggttcctt tgtcacatga
     1801 ggatcaccca tgtgaattgg tggctggtgt ggctaatgcc ctatggccct gaaaaagggc
     1861 cactggagga tattcatgca tctagccaaa ggttcctttg tcacatgagg atcacccatg
     1921 tgaattggtg gctggtgtgg ctaatgccct atggccctga aaaagggcca ctggaggata
     1981 ttcatgcatc tagagtcggg gtttaaacct agagcggccg ctggccgcaa taaaatatct
     2041 ttattttcat tacatctgtg tgttggtttt ttgtgtgagg atctaaatga gtcttcggac
     2101 ctcgcggggg ccgcttaagc ggtggttagg gtttgtctga cgcgggggga gggggaagga
     2161 acgaaacact ctcattcgga ggcggctcgg ggtttggtct tggtggccac gggcacgcag
     2221 aagagcgccg cgatcctctt aagcaccccc ccgccctccg tggaggcggg ggtttggtcg
     2281 gcgggtggta actggcgggc cgctgactcg ggcgggtcgc gcgccccaga gtgtgacctt
     2341 ttcggtctgc tcgcagaccc ccgggcggcg ccgccgcggc ggcgacgggc tcgctgggtc
     2401 ctaggctcca tggggaccgt atacgtggac aggctctgga gcatccgcac gactgcggtg
     2461 atattaccgg agaccttctg cgggacgagc cgggtcacgc ggctgacgcg gagcgtccgt
     2521 tgggcgacaa acaccaggac ggggcacagg tacactatct tgtcacccgg aggcgcgagg
     2581 gactgcagga gcttcaggga gtggcgcagc tgcttcatcc ccgtggcccg ttgctcgcgt
     2641 ttgctggcgg tgtccccgga agaaatatat ttgcatgtct ttagttctat gatgacacaa
     2701 accccgccca gcgtcttgtc attggcgaat tcgaacacgc agatgcagtc ggggcggcgc
     2761 ggtcccaggt ccacttcgca tattaaggtg acgcgtgtgg cctcgaacac cgagcgaccc
     2821 tgcagcgacc cgcttaaaag cttgattctt ctgacacaac agtctcgaac ttaagctgca
     2881 gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag
     2941 accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta
     3001 ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt
     3061 acagctctta aggctagagt acttaatacg actcactata ggctagcaaa gccaccatgg
     3121 ccgatgctaa gaacattaag aagggccctg ctcccttcta ccctctggag gatggcaccg
     3181 ctggcgagca gctgcacaag gccatgaaga ggtatgccct ggtgcctggc accattgcct
     3241 tcaccgatgc ccacattgag gtggacatca cctatgccga gtacttcgag atgtctgtgc
     3301 gcctggccga ggccatgaag aggtacggcc tgaacaccaa ccaccgcatc gtggtgtgct
     3361 ctgagaactc tctgcagttc ttcatgccag tgctgggcgc cctgttcatc ggagtggccg
     3421 tggcccctgc taacgacatt tacaacgagc gcgagctgct gaacagcatg ggcatttctc
     3481 agcctaccgt ggtgttcgtg tctaagaagg gcctgcagaa gatcctgaac gtgcagaaga
     3541 agctgcctat catccagaag atcatcatca tggactctaa gaccgactac cagggcttcc
     3601 agagcatgta cacattcgtg acatctcatc tgcctcctgg cttcaacgag tacgacttcg
     3661 tgccagagtc tttcgacagg gacaaaacca ttgccctgat catgaacagc tctgggtcta
     3721 ccggcctgcc taagggcgtg gccctgcctc atcgcaccgc ctgtgtgcgc ttctctcacg
     3781 cccgcgaccc tattttcggc aaccagatca tccccgacac cgctattctg agcgtggtgc
     3841 cattccacca cggcttcggc atgttcacca ccctgggcta cctgatttgc ggctttcggg
     3901 tggtgctgat gtaccgcttc gaggaggagc tgttcctgcg cagcctgcaa gactacaaaa
     3961 ttcagtctgc cctgctggtg ccaaccctgt tcagcttctt cgctaagagc accctgatcg
     4021 acaagtacga cctgtctaac ctgcacgaga ttgcctctgg cggcgcccca ctgtctaagg
     4081 aggtgggcga agccgtggcc aagcgctttc atctgccagg catccgccag ggctacggcc
     4141 tgaccgagac aaccagcgcc attctgatta ccccagaggg cgacgacaag cctggcgccg
     4201 tgggcaaggt ggtgccattc ttcgaggcca aggtggtgga cctggacacc ggcaagaccc
     4261 tgggagtgaa ccagcgcggc gagctgtgtg tgcgcggccc tatgattatg tccggctacg
     4321 tgaataaccc tgaggccaca aacgccctga tcgacaagga cggctggctg cactctggcg
     4381 acattgccta ctgggacgag gacgagcact tcttcatcgt ggaccgcctg aagtctctga
     4441 tcaagtacaa gggctaccag gtggccccag ccgagctgga gtctatcctg ctgcagcacc
     4501 ctaacatttt cgacgccgga gtggccggcc tgcccgacga cgatgccggc gagctgcctg
     4561 ccgccgtcgt cgtgctggaa cacggcaaga ccatgaccga gaaggagatc gtggactatg
     4621 tggccagcca ggtgacaacc gccaagaagc tgcgcggcgg agtggtgttc gtggacgagg
     4681 tgcccaaggg cctgaccggc aagctggacg cccgcaagat ccgcgagatc ctgatcaagg
     4741 ctaagaaagg cggcaagatc gccgtgtaat gcggccgctt cgagcagaca tgataagata
     4801 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     4861 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     4921 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     4981 caagtaaaac ctctacaaat gtggtaaaat cgataaggat ccaggtggca cttttcgggg
     5041 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     5101 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     5161 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     5221 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     5281 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     5341 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     5401 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     5461 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     5521 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     5581 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     5641 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5701 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     5761 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     5821 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     5881 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     5941 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6001 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6061 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     6121 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     6181 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     6241 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     6301 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     6361 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     6421 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     6481 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     6541 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     6601 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     6661 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     6721 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     6781 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tggct
//
