LOCUS       sequence-214321-        9230 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses wild type TPI reporter; contains partial TNFalpha 3' UTR; 
            enables detection of 5'-3' decay intermediates (xrFrag) 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 9230)
  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 9230)
  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-TNFalpha_3UTR-xrRNA-4H
FEATURES             Location/Qualifiers
     source          1..9230
                     /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    4934..5055
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4945..4964)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4999..5018
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      5241..5696
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5328..5347)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5538..5559
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5813..6170
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        5813..6004
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5836..5856)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6021..6156
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6221..7015
                     /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(6275..6294)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6885..6904
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7079..7127
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(7168..7187)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7287..7309
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7347..7365)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7433..7537
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7538..7606,7607..8398)
                     /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(7756..7775)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8569..9157
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9058..9077
                     /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 cttgagaaga cctcacctag aaattgacac aagtggacct taggccttcc tctctccaga
     3781 tgtttccaga cttccttgag acacggagcc cagccctccc catggagcca gctccctcta
     3841 tttatgtttg cacttgtgat tatttattat ttatttatta tttatttatt tacagatgaa
     3901 tgtatttatt tgggagaccg gggtatcctg ggggacccaa tgtaggagct gccttggctc
     3961 agacatgttt tccgtgaaaa cggagctgaa caataggctg ttcccatgta gccccctggc
     4021 ctctgtgcct tcttttgatt atgtttttta aaatatttat ctgattaagt tgtctaaaca
     4081 atgctgattt ggtgaccaac tgtcactcat tgctgagcct ctgctcccca ggggagttgt
     4141 gtctgtaatc gccctactag tcgaggtgag caagggcgag gagctgttca ccggggtggt
     4201 gcccatcctg gtggacttga tagtcaggcc agccggttag gctgccaccg aaggttggta
     4261 gacggtgctg cctgcgacca accccaggag gactgggtta ccaaagctga ttctccacgg
     4321 ttggaaagcc tcccagaacc gtctcggaag aggagtccct gccaacaatg gagatgaagc
     4381 ccgtgtcaga tcgcgaaagc gccacttcgc cgaggagtgc aatctgtgag gccccaggag
     4441 gactgggtaa acagtcgagc accccaccag tgcaggctgc ctatcagaaa gtggtggctg
     4501 gtgtggctaa tgccctggcc cacaagtatc actaagctcg ctttcttgct gtccaatttg
     4561 tcgagcaccc caccagtgca ggctgcctat cagaaagtgg tggctggtgt ggctaatgcc
     4621 ctggcccaca agtatcacta agctcgcttt cttgctgtcc aatttgtcga gcaccccacc
     4681 agtgcaggct gcctatcaga aagtggtggc tggtgtggct aatgccctgg cccacaagta
     4741 tcactaagct cgctttcttg ctgtccaatt tgtcgagcac cccaccagtg caggctgcct
     4801 atcagaaagt ggtggctggt gtggctaatg ccctggccca caagtatcac taagctcgct
     4861 ttcttgctgt ccaatttgtc gacccgggcg gccgcttccc tttagtgagg gttaatgctt
     4921 cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     4981 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     5041 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     5101 atgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat ccgataagga
     5161 tcgatccggg ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     5221 gcagcctgaa tggcgaatgg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     5281 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     5341 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     5401 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     5461 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     5521 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     5581 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     5641 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttcctg
     5701 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg cggatctgcg
     5761 cagcaccatg gcctgaaata acctctgaaa gaggaacttg gttaggtacc ttctgaggcg
     5821 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     5881 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     5941 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     6001 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     6061 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     6121 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagcttgatt
     6181 cttctgacac aacagtctcg aacttaaggc tagagccacc atgattgaac aagatggatt
     6241 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     6301 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     6361 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     6421 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     6481 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     6541 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     6601 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     6661 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     6721 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     6781 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     6841 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     6901 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     6961 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     7021 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg atggccgcaa
     7081 taaaatatct ttattttcat tacatctgtg tgttggtttt ttgtgtgaat cgatagcgat
     7141 aaggatccgc gtatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     7201 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     7261 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     7321 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     7381 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     7441 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     7501 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     7561 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     7621 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     7681 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     7741 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     7801 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     7861 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     7921 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     7981 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     8041 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     8101 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     8161 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     8221 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     8281 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     8341 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     8401 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     8461 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     8521 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     8581 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     8641 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     8701 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     8761 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     8821 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     8881 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     8941 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     9001 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     9061 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     9121 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     9181 acggttcctg gccttttgct ggccttttgc tcacatggct cgacagatct
//
