LOCUS       sequence-225469-        3247 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite gateway vector for 5' tagging with mKate2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3247)
  AUTHORS   Hall TE, Martel N, Ariotti N, Xiong Z, Lo HP, Ferguson C, Rae J, Lim
            YW, Parton RG
  TITLE     In vivo cell biological screening identifies an endocytic capture 
            mechanism for T-tubule formation.
  JOURNAL   Nat Commun. 2020 Jul 24;11(1):3711. doi: 10.1038/s41467-020-17486-w.
  PUBMED    32709891
REFERENCE   2  (bases 1 to 3247)
  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: "2020-07-24"; volume: "11(1):3711. doi"; pages: " 
            10.1038/s41467-020-17486-w"
COMMENT     Sequence Label: pME-mKate2-NS
FEATURES             Location/Qualifiers
     source          1..3247
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..100
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      101..110
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             107..802
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     protein_bind    complement(804..903)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(920..939)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(921..939)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(944..960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(944..960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1062..1081)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1073..1882
                     /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(1140..1159)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2029..2617
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2518..2537
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2771..2788
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2947..2974)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3066..3152)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3201..3223
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3215..3232
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3216..3232
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 caaataatga ttttattttg actgatagtg acctgttcgt tgcaacaaat tgatgagcaa
       61 tgctttttta taatgccaac tttgtacaaa aaagcaggct gccaccatgg tgagcgagct
      121 gattaaggag aacatgcaca tgaagctgta catggagggc accgtgaaca accaccactt
      181 caagtgcaca tccgagggcg aaggcaagcc ctacgagggc acccagacca tgagaatcaa
      241 ggcggtcgag ggcggccctc tccccttcgc cttcgacatc ctggctacca gcttcatgta
      301 cggcagcaaa accttcatca accacaccca gggcatcccc gacttcttta agcagtcctt
      361 ccccgagggc ttcacatggg agagagtcac cacatacgaa gacgggggcg tgctgaccgc
      421 tacccaggac accagcctcc aggacggctg cctcatctac aacgtcaaga tcagaggggt
      481 gaacttccca tccaacggcc ctgtgatgca gaagaaaaca ctcggctggg aggcctccac
      541 cgagaccctg taccccgctg acggcggcct ggaaggcaga gccgacatgg ccctgaagct
      601 cgtgggcggg ggccacctga tctgcaactt gaagaccaca tacagatcca agaaacccgc
      661 taagaacctc aagatgcccg gcgtctacta tgtggacaga agactggaaa gaatcaagga
      721 ggccgacaaa gagacctacg tcgagcagca cgaggtggct gtggccagat actgcgacct
      781 ccctagcaaa ctggggcaca gagacccagc tttcttgtac aaagttggca ttataagaaa
      841 gcattgctta tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat
      901 ttgccatcca gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg
      961 gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc
     1021 atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     1081 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
     1141 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
     1201 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
     1261 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
     1321 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     1381 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     1441 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     1501 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     1561 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     1621 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     1681 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     1741 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     1801 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     1861 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat
     1921 tacgctgact tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc
     1981 gtcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     2041 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     2101 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     2161 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     2221 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     2281 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     2341 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     2401 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     2461 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     2521 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     2581 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     2641 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
     2701 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
     2761 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccaatac gcaaaccgcc
     2821 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
     2881 agcgggcagt gagcgcaacg caattaatac gcgtaccgct agccaggaag agtttgtaga
     2941 aacgcaaaaa ggccatccgt caggatggcc ttctgcttag tttgatgcct ggcagtttat
     3001 ggcgggcgtc ctgcccgcca ccctccgggc cgttgcttca caacgttcaa atccgctccc
     3061 ggcggatttg tcctactcag gagagcgttc accgacaaac aacagataaa acgaaaggcc
     3121 cagtcttccg actgagcctt tcgttttatt tgatgcctgg cagttcccta ctctcgcgtt
     3181 aacgctagca tggatgtttt cccagtcacg acgttgtaaa acgacggcca gtcttaagct
     3241 cgggccc
//
