LOCUS       sequence-223866-        3394 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite gateway vector for expression of mKate2 fused to the 5' 
            and 3' sequences from zebrafish folate receptor 1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3394)
  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 3394)
  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-GPI-pA
FEATURES             Location/Qualifiers
     source          1..3394
                     /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             173..865
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="VSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIKA
                     VEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMALK
                     LVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARYC
                     DLPSKLGHR"
     protein_bind    complement(951..1050)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1067..1086)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1068..1086)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1091..1107)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1091..1107)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1209..1228)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1220..2029
                     /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(1287..1306)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2176..2764
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2665..2684
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2918..2935
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3094..3121)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3213..3299)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3348..3370
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3362..3379
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3363..3379
                     /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 acgctttatt
      121 tcacagacgt ctgctgctgc tgctgattgt actgccggcc tgcagcctcg aggtgagcga
      181 gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtga acaaccacca
      241 cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat
      301 caaggcggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta ccagcttcat
      361 gtacggcagc aaaaccttca tcaaccacac ccagggcatc cccgacttct ttaagcagtc
      421 cttccccgag ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac
      481 cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca agatcagagg
      541 ggtgaacttc ccatccaacg gccctgtgat gcagaagaaa acactcggct gggaggcctc
      601 caccgagacc ctgtaccccg ctgacggcgg cctggaaggc agagccgaca tggccctgaa
      661 gctcgtgggc gggggccacc tgatctgcaa cttgaagacc acatacagat ccaagaaacc
      721 cgctaagaac ctcaagatgc ccggcgtcta ctatgtggac agaagactgg aaagaatcaa
      781 ggaggccgac aaagagacct acgtcgagca gcacgaggtg gctgtggcca gatactgcga
      841 cctccctagc aaactggggc acagaggatc cctgaagggc ggtacggaga cggtcgccat
      901 ggcaacaggt ctgcatctga tggcgctgct ggtgatgctg ctgcgctgac acccagcttt
      961 cttgtacaaa gttggcatta taagaaagca ttgcttatca atttgttgca acgaacaggt
     1021 cactatcagt caaaataaaa tcattatttg ccatccagct gatatcccct atagtgagtc
     1081 gtattacatg gtcatagctg tttcctggca gctctggccc gtgtctcaaa atctctgatg
     1141 ttacattgca caagataaaa atatatcatc atgaacaata aaactgtctg cttacataaa
     1201 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcgaggc cgcgattaaa
     1261 ttccaacatg gatgctgatt tatatgggta taaatgggct cgcgataatg tcgggcaatc
     1321 aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg ccagagttgt ttctgaaaca
     1381 tggcaaaggt agcgttgcca atgatgttac agatgagatg gtcagactaa actggctgac
     1441 ggaatttatg cctcttccga ccatcaagca ttttatccgt actcctgatg atgcatggtt
     1501 actcaccact gcgatccccg gaaaaacagc attccaggta ttagaagaat atcctgattc
     1561 aggtgaaaat attgttgatg cgctggcagt gttcctgcgc cggttgcatt cgattcctgt
     1621 ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc gctcaggcgc aatcacgaat
     1681 gaataacggt ttggttgatg cgagtgattt tgatgacgag cgtaatggct ggcctgttga
     1741 acaagtctgg aaagaaatgc ataaactttt gccattctca ccggattcag tcgtcactca
     1801 tggtgatttc tcacttgata accttatttt tgacgagggg aaattaatag gttgtattga
     1861 tgttggacga gtcggaatcg cagaccgata ccaggatctt gccatcctat ggaactgcct
     1921 cggtgagttt tctccttcat tacagaaacg gctttttcaa aaatatggta ttgataatcc
     1981 tgatatgaat aaattgcagt ttcatttgat gctcgatgag tttttctaat cagaattggt
     2041 taattggttg taacactggc agagcattac gctgacttga cgggacggcg caagctcatg
     2101 accaaaatcc cttaacgtga gttacgcgtc gttccactga gcgtcagacc ccgtagaaaa
     2161 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     2221 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     2281 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     2341 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     2401 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     2461 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     2521 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     2581 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     2641 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     2701 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     2761 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     2821 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     2881 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     2941 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     3001 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatacgcg
     3061 taccgctagc caggaagagt ttgtagaaac gcaaaaaggc catccgtcag gatggccttc
     3121 tgcttagttt gatgcctggc agtttatggc gggcgtcctg cccgccaccc tccgggccgt
     3181 tgcttcacaa cgttcaaatc cgctcccggc ggatttgtcc tactcaggag agcgttcacc
     3241 gacaaacaac agataaaacg aaaggcccag tcttccgact gagcctttcg ttttatttga
     3301 tgcctggcag ttccctactc tcgcgttaac gctagcatgg atgttttccc agtcacgacg
     3361 ttgtaaaacg acggccagtc ttaagctcgg gccc
//
