LOCUS       sequence-318020-        5698 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Luciferase reporter for ribosome pausing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5698)
  AUTHORS   Han P, Shichino Y, Schneider-Poetsch T, Mito M, Hashimoto S, Udagawa
            T, Kohno K, Yoshida M, Mishima Y, Inada T, Iwasaki S
  TITLE     Genome-wide Survey of Ribosome Collision.
  JOURNAL   Cell Rep. 2020 May 5;31(5):107610. doi: 
            10.1016/j.celrep.2020.107610.
  PUBMED    32375038
REFERENCE   2  (bases 1 to 5698)
  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 Rep.";
            date: "2020-05-5"; volume: "31(5):107610. doi"; pages: " 
            10.1016/j.celrep.2020.107610"
COMMENT     Sequence Label: psiCHECK2-2A-3xFLAG-XBP1u_pause-2A
FEATURES             Location/Qualifiers
     source          1..5698
                     /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"
     primer_bind     604..623
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        666..684
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             706..1638
                     /codon_start=1
                     /product="Renilla luciferase"
                     /label=hRluc
                     /note="human codon-optimized"
                     /translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     CDS             1648..1704
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1705..1770
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1903..1959
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             1960..3609
                     /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="ADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAH
                     IEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPA
                     NDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSM
                     YTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHAR
                     DPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKI
                     QSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYG
                     LTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMSG
                     YVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILL
                     QHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVF
                     VDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             2916..2924
                     /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    3653..3774
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3664..3683)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3718..3737
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3907..4011
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4012..4080,4081..4872)
                     /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(4230..4249)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5043..5631
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5532..5551
                     /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 taggtttcct tcctcttttg gcaacatggc ttccaaggtg
      721 tacgaccccg agcaacgcaa acgcatgatc actgggcctc agtggtgggc tcgctgcaag
      781 caaatgaacg tgctggactc cttcatcaac tactatgatt ccgagaagca cgccgagaac
      841 gccgtgattt ttctgcatgg taacgctgcc tccagctacc tgtggaggca cgtcgtgcct
      901 cacatcgagc ccgtggctag atgcatcatc cctgatctga tcggaatggg taagtccggc
      961 aagagcggga atggctcata tcgcctcctg gatcactaca agtacctcac cgcttggttc
     1021 gagctgctga accttccaaa gaaaatcatc tttgtgggcc acgactgggg ggcttgtctg
     1081 gcctttcact actcctacga gcaccaagac aagatcaagg ccatcgtcca tgctgagagt
     1141 gtcgtggacg tgatcgagtc ctgggacgag tggcctgaca tcgaggagga tatcgccctg
     1201 atcaagagcg aagagggcga gaaaatggtg cttgagaata acttcttcgt cgagaccatg
     1261 ctcccaagca agatcatgcg gaaactggag cctgaggagt tcgctgccta cctggagcca
     1321 ttcaaggaga agggcgaggt tagacggcct accctctcct ggcctcgcga gatccctctc
     1381 gttaagggag gcaagcccga cgtcgtccag attgtccgca actacaacgc ctaccttcgg
     1441 gccagcgacg atctgcctaa gatgttcatc gagtccgacc ctgggttctt ttccaacgct
     1501 attgtcgagg gagctaagaa gttccctaac accgagttcg tgaaggtgaa gggcctccac
     1561 ttcagccagg aggacgctcc agatgaaatg ggtaagtaca tcaagagctt cgtggagcgc
     1621 gtgctgaaga acgagcaggg aagcggagct actaacttca gcctgctgaa gcaggctgga
     1681 gacgtggagg agaaccctgg acctgactac aaagaccatg acggtgatta taaagatcat
     1741 gacatcgatt acaaggatga cgatgacaag gcctggagga gctcccagag gtctacccag
     1801 aaggacccag ttccttacca gcctcccttt ctctgtcagt ggggacgtca tcagccaagc
     1861 tggaagccat taatgaactc attcgttttg accggaagcg gagctactaa cttcagcctg
     1921 ctgaagcagg ctggagacgt ggaggagaac cctggacctg ccgatgctaa gaacattaag
     1981 aagggccctg ctcccttcta ccctctggag gatggcaccg ctggcgagca gctgcacaag
     2041 gccatgaaga ggtatgccct ggtgcctggc accattgcct tcaccgatgc ccacattgag
     2101 gtggacatca cctatgccga gtacttcgag atgtctgtgc gcctggccga ggccatgaag
     2161 aggtacggcc tgaacaccaa ccaccgcatc gtggtgtgct ctgagaactc tctgcagttc
     2221 ttcatgccag tgctgggcgc cctgttcatc ggagtggccg tggcccctgc taacgacatt
     2281 tacaacgagc gcgagctgct gaacagcatg ggcatttctc agcctaccgt ggtgttcgtg
     2341 tctaagaagg gcctgcagaa gatcctgaac gtgcagaaga agctgcctat catccagaag
     2401 atcatcatca tggactctaa gaccgactac cagggcttcc agagcatgta cacattcgtg
     2461 acatctcatc tgcctcctgg cttcaacgag tacgacttcg tgccagagtc tttcgacagg
     2521 gacaaaacca ttgccctgat catgaacagc tctgggtcta ccggcctgcc taagggcgtg
     2581 gccctgcctc atcgcaccgc ctgtgtgcgc ttctctcacg cccgcgaccc tattttcggc
     2641 aaccagatca tccccgacac cgctattctg agcgtggtgc cattccacca cggcttcggc
     2701 atgttcacca ccctgggcta cctgatttgc ggctttcggg tggtgctgat gtaccgcttc
     2761 gaggaggagc tgttcctgcg cagcctgcaa gactacaaaa ttcagtctgc cctgctggtg
     2821 ccaaccctgt tcagcttctt cgctaagagc accctgatcg acaagtacga cctgtctaac
     2881 ctgcacgaga ttgcctctgg cggcgcccca ctgtctaagg aggtgggcga agccgtggcc
     2941 aagcgctttc atctgccagg catccgccag ggctacggcc tgaccgagac aaccagcgcc
     3001 attctgatta ccccagaggg cgacgacaag cctggcgccg tgggcaaggt ggtgccattc
     3061 ttcgaggcca aggtggtgga cctggacacc ggcaagaccc tgggagtgaa ccagcgcggc
     3121 gagctgtgtg tgcgcggccc tatgattatg tccggctacg tgaataaccc tgaggccaca
     3181 aacgccctga tcgacaagga cggctggctg cactctggcg acattgccta ctgggacgag
     3241 gacgagcact tcttcatcgt ggaccgcctg aagtctctga tcaagtacaa gggctaccag
     3301 gtggccccag ccgagctgga gtctatcctg ctgcagcacc ctaacatttt cgacgccgga
     3361 gtggccggcc tgcccgacga cgatgccggc gagctgcctg ccgccgtcgt cgtgctggaa
     3421 cacggcaaga ccatgaccga gaaggagatc gtggactatg tggccagcca ggtgacaacc
     3481 gccaagaagc tgcgcggcgg agtggtgttc gtggacgagg tgcccaaggg cctgaccggc
     3541 aagctggacg cccgcaagat ccgcgagatc ctgatcaagg ctaagaaagg cggcaagatc
     3601 gccgtgtaat aattctagag tcggggcggc cggccgcttc gagcagacat gataagatac
     3661 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
     3721 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
     3781 aacaattgca ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc
     3841 aagtaaaacc tctacaaatg tggtaaaatc gataaggatc caggtggcac ttttcgggga
     3901 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3961 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     4021 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     4081 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     4141 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     4201 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     4261 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     4321 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     4381 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     4441 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     4501 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     4561 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     4621 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     4681 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     4741 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     4801 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     4861 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     4921 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     4981 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     5041 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     5101 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     5161 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     5221 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     5281 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     5341 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     5401 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     5461 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     5521 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     5581 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     5641 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat ggctcgac
//
