LOCUS       sequence-209436-        8965 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses wild type TPI reporter; enables detection of 5'-3' decay 
            intermediates (xrFrag); 4MS2 tethering sites upstream of xrRNA and 
            allows detection via northern blot (4H binding sites).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8965)
  AUTHORS   Boehm V, Gerbracht JV, Marx MC, Gehring NH
  TITLE     Interrogating the degradation pathways of unstable mRNAs with 
            XRN1-resistant sequences.
  JOURNAL   Nat Commun. 2016 Dec 5;7:13691. doi: 10.1038/ncomms13691.
  PUBMED    27917860
REFERENCE   2  (bases 1 to 8965)
  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: "Nat 
            Commun."; date: "2016-12-5"; volume: "7:13691. doi"; pages: " 
            10.1038/ncomms13691"
COMMENT     Sequence Label: pCI-TPI_WT-4MS2-xrRNA-4H
FEATURES             Location/Qualifiers
     source          1..8965
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     680..700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     726..750
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          890..1022
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1005..1024
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1067..1085
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    4669..4790
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4680..4699)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4734..4753
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      4976..5431
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5063..5082)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5273..5294
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5548..5905
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5548..5739
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5571..5591)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5756..5891
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5956..6750
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6010..6029)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6620..6639
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6814..6862
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(6903..6922)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7022..7044
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7082..7100)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7168..7272
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7273..7341,7342..8133)
                     /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(7491..7510)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8304..8892
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8793..8812
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactg
      661 cgatcgcccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      721 agcagagctc gtttagtgaa ccgtcagatc actagaagct ttattgcggt agtttatcac
      781 agttaaattg ctaacgcagt cagtgcttct gacacaacag tctcgaactt aagctgcagt
      841 gactctctta aggtagcctt gcagaagttg gtcgtgaggc actgggcagg taagtatcaa
      901 ggttacaaga caggtttaag gagaccaata gaaactgggc ttgtcgagac agagaagact
      961 cttgcgtttc tgataggcac ctattggtct tactgacatc cactttgcct ttctctccac
     1021 aggtgtccac tcccagttca attacagctc ttaaggctag agtacttaat acgactcact
     1081 ataggctagc gcctcggctc cagcaccatg gcgccctcca ggaagttctt cgttggggga
     1141 aactggaaga tgaacgggcg gaagcagagt ctgggggagc tcatcggcac tctgaacgcg
     1201 gccaaggtgc cggccgacac cggtaagccc tcgccgagga ggggtctggc cgggccgggg
     1261 ccggggccgg ggcaggagtg gcagcgccct ctcccgaggc cccgaggccc cgaggccggt
     1321 atccgcgcgg acctgatgca gggctgtggg acgagggccg ctggggtccg ggcaggggcc
     1381 tcgcagccgc agccccgtcg gtgcgtcgag ggggcagggc ggagcacatg atgccccttg
     1441 gactatgggg caggtaagga cgttttcttc ccttcggcaa cctgaacgac tcccgccttc
     1501 cacggaaggg accgagcccg tgccaaacag gctgagcgat ttgggagtga ggagccatcc
     1561 taccgctttc cccaacctgg aaacagcaaa gcgcaaggcc tctgagtcag ttaggtctct
     1621 gccacccacg ggcaaaggat gctctcctcc atcctccttc ctccctccac cgaaatcgga
     1681 gagccgcggg cctgatccaa agaggcatcc ccttctcgtt cattccccag aggcctcaat
     1741 acaaacccca ggagttggcc cctctccttt tgctacaaat ccttgccttg caaaggggag
     1801 gtgaggatgg gctattttag aagggaagca gggttgctcc ctggagaatg ctgagtctgt
     1861 gaggtgccta tgccgagaat aggtcgagga aattggagcc ccagctgtta aaagagcaga
     1921 gggcagggtg agggccgtgg cctctcaggg gtatctggaa ggctcttcga gttgagtgca
     1981 gacccagcct gggctggaaa atggacaaag gtcatcttgc tggggtgaaa agggggagag
     2041 cagaaccaag aagaagaggg tgagggctgg ggggctccag ggcactggtt aggaattgtg
     2101 gggaatgaag gctttcttta gtctcatccc cctgtggtac catcttgtcc tcagaggtgg
     2161 tttgtgctcc ccctactgcc tatatcgact tcgcccggca gaagctagat cccaagattg
     2221 ctgtggctgc gcagaactgc tacaaagtga ctaatggggc ttttactggg gagatcaggt
     2281 gagatcgagg tggagagggg tgtgtgggac ccttccctca ctttcctcgt tgaggggaaa
     2341 gccacagggt gggctccctg ctgaaccttg gcttcatctc ttcctttagc cctggcatga
     2401 tcaaagactg cggagccacg tgggtggtcc tggggcactc agagagaagg catgtctttg
     2461 gggagtcaga tgaggttagt agccaagaga gaagataagg gatgtctttt tccaagaagg
     2521 atgtctcacc aagtctgttt ctcaacagct gattgggcag aaagtggccc atgctctggc
     2581 agagggactc ggagtaatcg cctgcattgg ggagaagcta gatgaaaggg aagctggcat
     2641 cactgagaag gttgttttcg agcagacaaa ggtcatcgca ggtatctctg gagaaaggga
     2701 cctttgagcc tatccagggc cacagagact cagagggtag ggtcaggccc tggagcctgt
     2761 cttggtcccc atgctgatcc agaaaaggaa aaaggggagg gggagtgaca atctttgctt
     2821 ggggcctatg acttctccag ccccaaggta gatgccacct ggaaatcccc caatgtccac
     2881 tagggggcag taggccaccg ttcttcgtac tccggagaac ctggctggag agctctttct
     2941 tgttcaccct tccctccatc tgtatctctg ccctgcagat aacgtgaagg actggagcaa
     3001 ggtcgtcctg gcctatgagc ctgtgtgggc cattggtact ggcaagactg caacacccca
     3061 acaggtaacc gggcccagga gccctgccct catcccagcc tgcctcaata ggtttggaca
     3121 gacacagccc acatggggca accccttatt tcaaagacac agagaccttg aacccagaga
     3181 cagtgacttg tccaagggca tccagtccag ggcctggctt ggatcagagc cctggtactc
     3241 tgactcagtc agaaaccaca ctaagtgtcc actggtgcca gtgatttttc ctcttagaga
     3301 ggcagaaaag gtcttactta ggccagcttc ttgttctagg cccaggaagt acacgagaag
     3361 ctccgaggat ggctgaagtc caacgtctct gatgcggtgg ctcagagcac ccgtatcatt
     3421 tatggaggtg agtggctttg gttcccggct gaggtggagt gggctgagga ctagactgag
     3481 ccctcggaca tggaggtggg gatggggcag actcatccca ttcttgacca agcccttgtt
     3541 ctgctccctt cccaggctct gtgactgggg caacctgcaa ggagctggcc agccagcctg
     3601 atgtggatgg cttccttgtg ggtggtgctt ccctcaagcc cgaattcgtg gacatcatca
     3661 atgccaaaca atgagcccca tccatcttcc ctacccttcc tgccaagcca gggactcgag
     3721 tattgcctag aaagcacgag catcagccgt gcctccaggt cgaatcttca aacgacgacg
     3781 atcacgcgtc gctgcagtgc aggagtcgag tattgcctag aaagcacgag catcagccgt
     3841 gcctccaggt cgaatcttca aacgacgacg atcacgcgtc gctgcagtgc aggagtcgag
     3901 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtgga cttgatagtc
     3961 aggccagccg gttaggctgc caccgaaggt tggtagacgg tgctgcctgc gaccaacccc
     4021 aggaggactg ggttaccaaa gctgattctc cacggttgga aagcctccca gaaccgtctc
     4081 ggaagaggag tccctgccaa caatggagat gaagcccgtg tcagatcgcg aaagcgccac
     4141 ttcgccgagg agtgcaatct gtgaggcccc aggaggactg ggtaaacagt cgagcacccc
     4201 accagtgcag gctgcctatc agaaagtggt ggctggtgtg gctaatgccc tggcccacaa
     4261 gtatcactaa gctcgctttc ttgctgtcca atttgtcgag caccccacca gtgcaggctg
     4321 cctatcagaa agtggtggct ggtgtggcta atgccctggc ccacaagtat cactaagctc
     4381 gctttcttgc tgtccaattt gtcgagcacc ccaccagtgc aggctgccta tcagaaagtg
     4441 gtggctggtg tggctaatgc cctggcccac aagtatcact aagctcgctt tcttgctgtc
     4501 caatttgtcg agcaccccac cagtgcaggc tgcctatcag aaagtggtgg ctggtgtggc
     4561 taatgccctg gcccacaagt atcactaagc tcgctttctt gctgtccaat ttgtcgaccc
     4621 gggcggccgc ttccctttag tgagggttaa tgcttcgagc agacatgata agatacattg
     4681 atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt
     4741 gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca
     4801 attgcattca ttttatgttt caggttcagg gggagatgtg ggaggttttt taaagcaagt
     4861 aaaacctcta caaatgtggt aaaatccgat aaggatcgat ccgggctggc gtaatagcga
     4921 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg
     4981 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     5041 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     5101 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     5161 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     5221 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     5281 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     5341 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     5401 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     5461 tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca ccatggcctg aaataacctc
     5521 tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg
     5581 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     5641 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     5701 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     5761 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     5821 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     5881 ttttggaggc ctaggctttt gcaaaaagct tgattcttct gacacaacag tctcgaactt
     5941 aaggctagag ccaccatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     6001 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     6061 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     6121 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     6181 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     6241 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     6301 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     6361 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     6421 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     6481 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     6541 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     6601 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     6661 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     6721 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc
     6781 aagcgacgcc caacctgcca tcacgatggc cgcaataaaa tatctttatt ttcattacat
     6841 ctgtgtgttg gttttttgtg tgaatcgata gcgataagga tccgcgtatg gtgcactctc
     6901 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     6961 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     7021 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     7081 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     7141 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     7201 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     7261 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     7321 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     7381 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     7441 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     7501 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     7561 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     7621 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     7681 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     7741 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     7801 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     7861 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     7921 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     7981 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     8041 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     8101 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     8161 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     8221 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     8281 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     8341 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     8401 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     8461 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     8521 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     8581 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     8641 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     8701 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     8761 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     8821 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     8881 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     8941 tttgctcaca tggctcgaca gatct
//
