LOCUS       sequence-237078-        5005 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pMVP L5-L4 entry plasmid, contains Tir1-P2A-AID for 4-component 
            MultiSite Gateway Pro assembly. Allows expression of N-term Tir1 
            linked by P2A to AID domain fused to gene of interest..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5005)
  AUTHORS   Haldeman JM, Conway AE, Arlotto ME, Slentz DH, Muoio DM, Becker TC, 
            Newgard CB
  TITLE     Creation of versatile cloning platforms for transgene expression and
            dCas9-based epigenome editing.
  JOURNAL   Nucleic Acids Res. 2018 Dec 27. pii: 5264291. doi: 
            10.1093/nar/gky1286.
  PUBMED    30590691
REFERENCE   2  (bases 1 to 5005)
  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: "Nucleic 
            Acids Res. 2018 Dec 27. pii: 5264291. doi: 10.1093/nar/gky1286."
COMMENT     Sequence Label: JE301: pMVP (L5-L4) Tir1-P2A-AID
FEATURES             Location/Qualifiers
     source          1..5005
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    570..665
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      668..677
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             722..2443
                     /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             2471..2500
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2507..2536
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2543..2572
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2591..2620
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2627..2656
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2663..2692
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2711..2740
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2747..2776
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2783..2812
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2846..2902
                     /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             2903..3106
                     /codon_start=1
                     /product="minimal degron from the auxin-responsive IAA17 
                     protein of Arabidopsis thaliana (Kubota et al., 2013)"
                     /label=mini-AID
                     /note="acts as an auxin-inducible degron (AID) in the 
                     presence of TIR1"
                     /translation="KEKSACPKDPAKPPAKAQVVGWPPVRSYRKNVMVSCQKSSGGPEA
                     AAFVKVSMDGAPYLRKIDLRMYK"
     protein_bind    complement(3120..3215)
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(3246..3265)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3247..3265)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3270..3286)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3270..3286)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3388..3407)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3399..4208
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3466..3485)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4355..4943
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4844..4863
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtatacaaa
      661 agttgtcgcc accatggcat tacaaaaaaa attagaagaa ttagaattag atgaaggcgg
      721 cacgtacttc ccggaggagg tggtggagca catcttcagc ttcctgccgg cgcagcgcga
      781 ccgcaacacg gtctcgctcg tctgcaaggt gtggtacgag atcgagaggc tgagccgccg
      841 cggcgtcttc gtgggcaact gctacgccgt gcgcgccggc cgcgtcgccg cgcggttccc
      901 caacgtgcgg gcgctcacgg tgaaggggaa gccccacttc gccgacttca acctcgtgcc
      961 ccccgactgg ggcggctacg cggggccgtg gatcgaggcg gccgcgaggg gatgccacgg
     1021 cctggaggag ctcaggatga agcggatggt ggtgtccgac gagagcctcg agctgctggc
     1081 tcgctcgttc ccgcggttca gggctcttgt tcttatcagc tgcgaggggt tcagcactga
     1141 cgggctagcc gccgtcgcga gccattgcaa gcttctgagg gagttggatt tgcaggaaaa
     1201 tgaagtggag gatcgagggc ctaggtggct ttcctgcttc cctgattcct gcacatcact
     1261 tgtctcattg aattttgcct gcatcaaagg ggaggttaat gctggttcac tggagagact
     1321 tgttagcagg tccccaaacc tgcggagttt gaggctgaat cgatctgtat cggtagatac
     1381 acttgcaaag atactactgc gtacccctaa cttggaggat ttggggacag ggaatttgac
     1441 agatgacttc caaactgagt cctactttaa gcttaccagt gctctggaga aatgcaagat
     1501 gttgaggagt ttgtctggat tctgggatgc ttctcctgtt tgcctgtcat ttatctaccc
     1561 cctgtgtgct caactgacag gattgaactt gagctatgca cccacacttg atgcttctga
     1621 ccttacaaaa atgattagcc gctgtgtgaa gctccaacgc ctttgggtac tggattgtat
     1681 ctcggacaaa ggcttgcaag tggtggcctc cagttgcaaa gacttgcaag aactcagggt
     1741 atttccatca gatttctacg tagctggtta ttctgcagtg acagaggagg gacttgttgc
     1801 agtatccttg ggctgtccaa aactgaactc actactgtac ttctgtcacc aaatgactaa
     1861 tgctgcacta gttactgtcg ccaagaactg tccaaatttc acacgattca gactttgtat
     1921 tcttgagcca gggaagcctg atgttgtgac aagccaacca ttagatgaag gctttggagc
     1981 tattgttcgt gagtgcaagg gattacaacg tttgtcaata tctggtcttc tcacagacaa
     2041 agttttcatg tatatcggga aatatgcaaa acaacttgag atgctttcta tagcatttgc
     2101 tggtgacagt gataagggta tgatgcatgt tatgaatgga tgcaagaatt taaggaaact
     2161 ggagataaga gatagcccgt ttggtgatgc tgcactcttg gggaattttg ctaggtacga
     2221 gacaatgcga tccctttgga tgtcatcttg caatgtcacg ttaaaggggt gccaagtcct
     2281 tgcgtcaaag atgccgatgc tcaatgttga ggtcataaat gagcgggatg gtagcaatga
     2341 aatggaggaa aaccatggag atctgcctaa agtggagaaa ttatatgtgt accgcacaac
     2401 tgctggggcg agggatgatg caccaaattt tgttaaaatc ctagtcgact ccggttctgc
     2461 tgctagtggt gaacaaaagt tgatttctga agaagatttg aacggtgaac aaaagctaat
     2521 ctccgaggaa gacttgaacg gtgaacaaaa attaatctca gaagaagact tgaacggatc
     2581 ctctagaggt gaacaaaagt tgatttctga agaagatttg aacggtgaac aaaagctaat
     2641 ctccgaggaa gacttgaacg gtgaacaaaa attaatctca gaagaagact tgaacggatc
     2701 ctctagaggt gaacaaaagt tgatttctga agaagatttg aacggtgaac aaaagctaat
     2761 ctccgaggaa gacttgaacg gtgaacaaaa attaatctca gaagaagact tgaacggatc
     2821 cactagcggt acagccggat caggagctac taatttctct ctgttaaagc aggcaggaga
     2881 tgtggaggag aatccaggtc ctaaggagaa gagtgcttgt cctaaagatc cagccaaacc
     2941 tccggccaag gcacaagttg tgggatggcc accggtgaga tcataccgga agaacgtgat
     3001 ggtttcctgc caaaaatcaa gcggtggccc ggaggcggcg gcgttcgtga aggtatcaat
     3061 ggacggagca ccgtacttga ggaaaatcga tttgaggatg tataaaggca gtaccaaccc
     3121 aacttttcta tacaaagttg gcattataag aaagcattgc ttatcaattt gttgcaacga
     3181 acaggtcact atcagtcaaa ataaaatcat tatttgccat ccagctgcag ggcggccgcg
     3241 atatccccta tagtgagtcg tattacatgg tcatagctgt ttcctggcag ctctggcccg
     3301 tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa
     3361 aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa
     3421 cgtcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc
     3481 gcgataatgt cgggcaatca ggtgcgacaa tctatcgctt gtatgggaag cccgatgcgc
     3541 cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg
     3601 tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta
     3661 ctcctgatga tgcatggtta ctcaccactg cgatccccgg aaaaacagca ttccaggtat
     3721 tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc
     3781 ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg
     3841 ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc
     3901 gtaatggctg gcctgttgaa caagtctgga aagaaatgca taaacttttg ccattctcac
     3961 cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga
     4021 aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg
     4081 ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa
     4141 aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt
     4201 ttttctaatc agaattggtt aattggttgt aacactggca gagcattacg ctgacttgac
     4261 gggacggcgc aagctcatga ccaaaatccc ttaacgtgag ttacgcgtcg ttccactgag
     4321 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     4381 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     4441 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     4501 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     4561 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     4621 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     4681 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     4741 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     4801 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     4861 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     4921 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     4981 tttgctggcc ttttgctcac atgtt
//
