LOCUS       sequence-227023-        5888 bp    DNA     circular SYN 14-APR-2025
DEFINITION  mRNA synthesis of NLS-TIR1-HA for injection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5888)
  AUTHORS   Daniel K, Icha J, Horenburg C, Muller D, Norden C, Mansfeld J
  TITLE     Conditional control of fluorescent protein degradation by an 
            auxin-dependent nanobody.
  JOURNAL   Nat Commun. 2018 Aug 17;9(1):3297. doi: 10.1038/s41467-018-05855-5.
  PUBMED    30120238
REFERENCE   2  (bases 1 to 5888)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2018-08-17"; volume: "9(1):3297. doi"; pages: " 
            10.1038/s41467-018-05855-5"
COMMENT     Sequence Label: pCS2+_NLS-Tir1-HA
FEATURES             Location/Qualifiers
     source          1..5888
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        42..1019
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1054..1072
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1054..1071
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      1119..1128
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1140..1160
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1167..2888
                     /codon_start=1
                     /product="rice transport inhibitor response 1 protein"
                     /label=OsTIR1
                     /note="F-box protein that mediates auxin-dependent 
                     interaction of a ubiquitin ligase with an auxin-inducible 
                     degron (Tan et al., 2007)"
                     /translation="TYFPEEVVEHIFSFLPAQRDRNTVSLVCKVWYEIERLSRRGVFVG
                     NCYAVRAGRVAARFPNVRALTVKGKPHFADFNLVPPDWGGYAGPWIEAAARGCHGLEEL
                     RMKRMVVSDESLELLARSFPRFRALVLISCEGFSTDGLAAVASHCKLLRELDLQENEVE
                     DRGPRWLSCFPDSCTSLVSLNFACIKGEVNAGSLERLVSRSPNLRSLRLNRSVSVDTLA
                     KILLRTPNLEDLGTGNLTDDFQTESYFKLTSALEKCKMLRSLSGFWDASPVCLSFIYPL
                     CAQLTGLNLSYAPTLDASDLTKMISRCVKLQRLWVLDCISDKGLQVVASSCKDLQELRV
                     FPSDFYVAGYSAVTEEGLVAVSLGCPKLNSLLYFCHQMTNAALVTVAKNCPNFTRFRLC
                     ILEPGKPDVVTSQPLDEGFGAIVRECKGLQRLSISGLLTDKVFMYIGKYAKQLEMLSIA
                     FAGDSDKGMMHVMNGCKNLRKLEIRDSPFGDAALLGNFARYETMRSLWMSSCNVTLKGC
                     QVLASKMPMLNVEVINERDGSNEMEENHGDLPKVEKLYVYRTTAGARDDAPNFVKIL"
     CDS             2892..2918
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    2959..3093
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2983..3002)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3037..3056
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(3206..3226)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3206..3224)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3245..3261)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3245..3261)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3258..3280)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3269..3285
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3293..3299,3300..3317,3318..3323))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3338..3359
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3476..3493)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3647..4235)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3727..3746)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4406..5197,5198..5266))
                     /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     5029..5048
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5267..5371)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5397..5852)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5534..5555)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5746..5765
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ggcgatcggt gcgggcctct tcgctattac gccagtcgac cgccaattca atatggcgta
       61 tatggactca tgccaattca atatggtgga tctggacctg tgccaattca atatggcgta
      121 tatggactcg tgccaattca atatggtgga tctggacccc agccaattca atatggcgga
      181 cttggcacca tgccaattca atatggcgga cctggcactg tgccaactgg ggaggggtct
      241 acttggcacg gtgccaagtt tgaggagggg tcttggccct gtgccaagtc cgccatattg
      301 aattggcatg gtgccaataa tggcggccat attggctata tgccaggatc aatatatagg
      361 caatatccaa tatggcccta tgccaatatg gctattggcc aggttcaata ctatgtattg
      421 gccctatgcc atatagtatt ccatatatgg gttttcctat tgacgtagat agcccctccc
      481 aatgggcggt cccatatacc atatatgggg cttcctaata ccgcccatag ccactccccc
      541 attgacgtca atggtctcta tatatggtct ttcctattga cgtcatatgg gcggtcctat
      601 tgacgtatat ggcgcctccc ccattgacgt caattacggt aaatggcccg cctggctcaa
      661 tgcccattga cgtcaatagg accacccacc attgacgtca atgggatggc tcattgccca
      721 ttcatatccg ttctcacgcc ccctattgac gtcaatgacg gtaaatggcc cacttggcag
      781 tacatcaata tctattaata gtaacttggc aagtacatta ctattggaag tacgccaggg
      841 tacattggca gtactcccat tgacgtcaat ggcggtaaat ggcccgcgat ggctgccaag
      901 tacatcccca ttgacgtcaa tggggagggg caatgacgca aatgggcgtt ccattgacgt
      961 aaatgggcgg taggcgtgcc taatgggagg tctatataag caatgctcgt ttagggaacc
     1021 gccattctgc ctggggacgt cggagcaagc ttgatttagg tgacactata gaatacaagc
     1081 tacttgttct ttttgcagga tcccatcgat tacccgccgc caccatgggc cccgccgccc
     1141 ccaagaagaa gagaaaggtg ctggacacgt acttccctga agaagttgtg gaacatatct
     1201 tttcttttct tcctgcccaa agagaccgga atacagtctc tttggtatgt aaggtttggt
     1261 atgagattga aaggttgtca aggagaggag tgtttgtggg aaactgctat gcggtaaggg
     1321 ctggaagagt cgctgccaga ttccccaatg tgagggcact tacagttaag ggtaaaccac
     1381 attttgctga tttcaatttg gttcctccgg actggggagg ttatgctgga ccatggattg
     1441 aggctgctgc cagaggttgt catgggttgg aggagttgag gatgaagagg atggttgtgt
     1501 ctgatgagtc ccttgaactt ctggcccggt ccttccccag attcagagct ttggtcttga
     1561 tttcttgtga gggtttctct actgacggac tcgctgcagt cgcttcccac tgtaagttgt
     1621 tgagggagtt ggacctccaa gaaaatgagg ttgaggacag aggaccaaga tggttgtcat
     1681 gtttccctga ctcatgtact tcccttgttt ccttgaattt cgcgtgtatc aaaggagagg
     1741 tcaatgccgg gagtcttgag agattggtct caagaagtcc caacttgcga tcactgaggc
     1801 tcaacagatc tgtgagtgtg gataccttgg caaagattct cttgagaacc cccaatttgg
     1861 aggaccttgg aactgggaat ctcacggatg acttccaaac tgagtcttac tttaagttga
     1921 ctagtgcttt ggaaaagtgt aagatgttga gatcactgtc cggattctgg gatgcttcac
     1981 ctgtttgtct ttccttcatc taccctcttt gtgcccagct gactggactc aacttgtcct
     2041 atgctccaac actggatgcg agtgacttga ccaagatgat ttcaagatgc gtgaagcttc
     2101 aaagactttg ggtcctcgat tgtatctccg acaaaggact tcaagtggtg gcatcatctt
     2161 gtaaggatct ccaagagttg agagtgtttc caagtgactt ctatgtggca ggttacagtg
     2221 ctgtaactga ggaagggttg gttgctgtct cccttggatg tcccaagttg aattcactct
     2281 tgtacttttg tcaccaaatg actaacgctg cccttgtgac agtggccaaa aattgcccta
     2341 acttcacacg cttcagactt tgtatccttg aacctgggaa acctgacgtg gtcaccagtc
     2401 agcccttgga cgaaggattt ggggcgattg tgagagaatg taagggactc caaagactct
     2461 ctatcagtgg actcctgact gacaaggtgt ttatgtacat tggtaagtac gctaagcagc
     2521 tggaaatgct gtctattgcc tttgccggtg attctgacaa ggggatgatg catgttatga
     2581 atggatgtaa gaacttgaga aagcttgaga taagagacag tccattcgga gatgcagccc
     2641 tccttggtaa cttcgcaaga tacgaaacta tgaggtctct ttggatgtcc agctgtaatg
     2701 ttacactcaa gggatgtcaa gtgctggcct caaaaatgcc aatgctgaat gtggaggtga
     2761 tcaacgagag agatggatcc aatgaaatgg aagagaacca cggagatctt cccaaggtgg
     2821 aaaagttgta tgtctacaga accactgctg gggcaaggga tgacgctcca aacttcgtga
     2881 aaatactggg ctacccatac gatgttcctg actatgcgtg actcgagcct ctagaactat
     2941 agtgagtcgt attacgtaga tccagacatg ataagataca ttgatgagtt tggacaaacc
     3001 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     3061 tttgtaacca ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg
     3121 tttcaggttc agggggaggt gtgggaggtt ttttaattcg cggccgcggc gccaatgcat
     3181 tgggcccggt acccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat
     3241 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3301 gagccgggag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3361 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     3421 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     3481 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     3541 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     3601 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     3661 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     3721 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     3781 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     3841 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     3901 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     3961 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4021 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4081 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4141 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4201 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4261 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4321 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4381 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     4441 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     4501 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     4561 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     4621 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     4681 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     4741 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     4801 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     4861 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     4921 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     4981 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5041 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5101 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5161 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5221 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5281 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5341 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat
     5401 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     5461 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     5521 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     5581 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     5641 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     5701 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     5761 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     5821 cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg
     5881 ttgggaag
//
