LOCUS       sequence-263060-        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        62..419
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        62..253
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(85..105)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      270..405
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          489..621
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      688..697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             694..1629
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     misc_RNA        1683..1701
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        1791..1809
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        1899..1917
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     polyA_signal    2025..2073
                     /note="synthetic polyadenylation signal"
     promoter        2082..2833
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          2903..3035
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     promoter        3080..3098
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      3107..3116
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3113..4765
                     /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             4072..4080
                     /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    4790..4911
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4801..4820)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4855..4874
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        5044..5148
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5149..5217,5218..6009)
                     /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(5367..5386)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6180..6768
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6669..6688
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 agatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttaggtacct
       61 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
      121 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
      181 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
      241 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
      301 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
      361 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
      421 agcttgattc ttctgacaca acagtctcga acttaagctg cagaagttgg tcgtgaggca
      481 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      541 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      601 actttgcctt tctctccaca ggtgtccact cccagttcaa ttacagctct taaggctaga
      661 gtacttaata cgactcacta taggctagcc accatggctt ccaaggtgta cgaccccgag
      721 caacgcaaac gcatgatcac tgggcctcag tggtgggctc gctgcaagca aatgaacgtg
      781 ctggactcct tcatcaacta ctatgattcc gagaagcacg ccgagaacgc cgtgattttt
      841 ctgcatggta acgctgcctc cagctacctg tggaggcacg tcgtgcctca catcgagccc
      901 gtggctagat gcatcatccc tgatctgatc ggaatgggta agtccggcaa gagcgggaat
      961 ggctcatatc gcctcctgga tcactacaag tacctcaccg cttggttcga gctgctgaac
     1021 cttccaaaga aaatcatctt tgtgggccac gactgggggg cttgtctggc ctttcactac
     1081 tcctacgagc accaagacaa gatcaaggcc atcgtccatg ctgagagtgt cgtggacgtg
     1141 atcgagtcct gggacgagtg gcctgacatc gaggaggata tcgccctgat caagagcgaa
     1201 gagggcgaga aaatggtgct tgagaataac ttcttcgtcg agaccatgct cccaagcaag
     1261 atcatgcgga aactggagcc tgaggagttc gctgcctacc tggagccatt caaggagaag
     1321 ggcgaggtta gacggcctac cctctcctgg cctcgcgaga tccctctcgt taagggaggc
     1381 aagcccgacg tcgtccagat tgtccgcaac tacaacgcct accttcgggc cagcgacgat
     1441 ctgcctaaga tgttcatcga gtccgaccct gggttctttt ccaacgctat tgtcgaggga
     1501 gctaagaagt tccctaacac cgagttcgtg aaggtgaagg gcctccactt cagccaggag
     1561 gacgctccag atgaaatggg taagtacatc aagagcttcg tggagcgcgt gctgaagaac
     1621 gagcagtaat tctaggcgat cgctcgagcc gtgtaataat tctagccaaa ggttcctttg
     1681 tcacatgagg atcacccatg tgaattggtg gctggtgtgg ctaatgccct atggccctga
     1741 aaaagggcca ctggaggata ttcatgcatc tagccaaagg ttcctttgtc acatgaggat
     1801 cacccatgtg aattggtggc tggtgtggct aatgccctat ggccctgaaa aagggccact
     1861 ggaggatatt catgcatcta gccaaaggtt cctttgtcac atgaggatca cccatgtgaa
     1921 ttggtggctg gtgtggctaa tgccctatgg ccctgaaaaa gggccactgg aggatattca
     1981 tgcatctaga gtcggggttt aaacctagag cggccgctgg ccgcaataaa atatctttat
     2041 tttcattaca tctgtgtgtt ggttttttgt gtgaggatct aaatgagtct tcggacctcg
     2101 cgggggccgc ttaagcggtg gttagggttt gtctgacgcg gggggagggg gaaggaacga
     2161 aacactctca ttcggaggcg gctcggggtt tggtcttggt ggccacgggc acgcagaaga
     2221 gcgccgcgat cctcttaagc acccccccgc cctccgtgga ggcgggggtt tggtcggcgg
     2281 gtggtaactg gcgggccgct gactcgggcg ggtcgcgcgc cccagagtgt gaccttttcg
     2341 gtctgctcgc agacccccgg gcggcgccgc cgcggcggcg acgggctcgc tgggtcctag
     2401 gctccatggg gaccgtatac gtggacaggc tctggagcat ccgcacgact gcggtgatat
     2461 taccggagac cttctgcggg acgagccggg tcacgcggct gacgcggagc gtccgttggg
     2521 cgacaaacac caggacgggg cacaggtaca ctatcttgtc acccggaggc gcgagggact
     2581 gcaggagctt cagggagtgg cgcagctgct tcatccccgt ggcccgttgc tcgcgtttgc
     2641 tggcggtgtc cccggaagaa atatatttgc atgtctttag ttctatgatg acacaaaccc
     2701 cgcccagcgt cttgtcattg gcgaattcga acacgcagat gcagtcgggg cggcgcggtc
     2761 ccaggtccac ttcgcatatt aaggtgacgc gtgtggcctc gaacaccgag cgaccctgca
     2821 gcgacccgct taaaagcttg attcttctga cacaacagtc tcgaacttaa gctgcagaag
     2881 ttggtcgtga ggcactgggc aggtaagtat caaggttaca agacaggttt aaggagacca
     2941 atagaaactg ggcttgtcga gacagagaag actcttgcgt ttctgatagg cacctattgg
     3001 tcttactgac atccactttg cctttctctc cacaggtgtc cactcccagt tcaattacag
     3061 ctcttaaggc tagagtactt aatacgactc actataggct agcaaagcca ccatggccga
     3121 tgctaagaac attaagaagg gccctgctcc cttctaccct ctggaggatg gcaccgctgg
     3181 cgagcagctg cacaaggcca tgaagaggta tgccctggtg cctggcacca ttgccttcac
     3241 cgatgcccac attgaggtgg acatcaccta tgccgagtac ttcgagatgt ctgtgcgcct
     3301 ggccgaggcc atgaagaggt acggcctgaa caccaaccac cgcatcgtgg tgtgctctga
     3361 gaactctctg cagttcttca tgccagtgct gggcgccctg ttcatcggag tggccgtggc
     3421 ccctgctaac gacatttaca acgagcgcga gctgctgaac agcatgggca tttctcagcc
     3481 taccgtggtg ttcgtgtcta agaagggcct gcagaagatc ctgaacgtgc agaagaagct
     3541 gcctatcatc cagaagatca tcatcatgga ctctaagacc gactaccagg gcttccagag
     3601 catgtacaca ttcgtgacat ctcatctgcc tcctggcttc aacgagtacg acttcgtgcc
     3661 agagtctttc gacagggaca aaaccattgc cctgatcatg aacagctctg ggtctaccgg
     3721 cctgcctaag ggcgtggccc tgcctcatcg caccgcctgt gtgcgcttct ctcacgcccg
     3781 cgaccctatt ttcggcaacc agatcatccc cgacaccgct attctgagcg tggtgccatt
     3841 ccaccacggc ttcggcatgt tcaccaccct gggctacctg atttgcggct ttcgggtggt
     3901 gctgatgtac cgcttcgagg aggagctgtt cctgcgcagc ctgcaagact acaaaattca
     3961 gtctgccctg ctggtgccaa ccctgttcag cttcttcgct aagagcaccc tgatcgacaa
     4021 gtacgacctg tctaacctgc acgagattgc ctctggcggc gccccactgt ctaaggaggt
     4081 gggcgaagcc gtggccaagc gctttcatct gccaggcatc cgccagggct acggcctgac
     4141 cgagacaacc agcgccattc tgattacccc agagggcgac gacaagcctg gcgccgtggg
     4201 caaggtggtg ccattcttcg aggccaaggt ggtggacctg gacaccggca agaccctggg
     4261 agtgaaccag cgcggcgagc tgtgtgtgcg cggccctatg attatgtccg gctacgtgaa
     4321 taaccctgag gccacaaacg ccctgatcga caaggacggc tggctgcact ctggcgacat
     4381 tgcctactgg gacgaggacg agcacttctt catcgtggac cgcctgaagt ctctgatcaa
     4441 gtacaagggc taccaggtgg ccccagccga gctggagtct atcctgctgc agcaccctaa
     4501 cattttcgac gccggagtgg ccggcctgcc cgacgacgat gccggcgagc tgcctgccgc
     4561 cgtcgtcgtg ctggaacacg gcaagaccat gaccgagaag gagatcgtgg actatgtggc
     4621 cagccaggtg acaaccgcca agaagctgcg cggcggagtg gtgttcgtgg acgaggtgcc
     4681 caagggcctg accggcaagc tggacgcccg caagatccgc gagatcctga tcaaggctaa
     4741 gaaaggcggc aagatcgccg tgtaatgcgg ccgcttcgag cagacatgat aagatacatt
     4801 gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt
     4861 tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac
     4921 aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag
     4981 taaaacctct acaaatgtgg taaaatcgat aaggatccag gtggcacttt tcggggaaat
     5041 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     5101 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     5161 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     5221 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     5281 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     5341 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     5401 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     5461 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     5521 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     5581 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     5641 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     5701 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     5761 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     5821 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     5881 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     5941 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     6001 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     6061 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     6121 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     6181 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     6241 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     6301 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     6361 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     6421 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     6481 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     6541 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     6601 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     6661 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     6721 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     6781 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatggc tcgac
//
