LOCUS       sequence-223868-        3321 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite gateway vector for expression of mKate2 fused to the 5' 
            signal sequence from calreticulin and 3' KDEL ER retention sequence 
            (marks the ER, or ER/SR in muscle).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3321)
  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 3321)
  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-KDEL-pA
FEATURES             Location/Qualifiers
     source          1..3321
                     /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"
     CDS             164..856
                     /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(878..977)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(994..1013)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(995..1013)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1018..1034)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1018..1034)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1136..1155)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1147..1956
                     /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(1214..1233)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2103..2691
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2592..2611
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2845..2862
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3021..3048)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3140..3226)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3275..3297
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3289..3306
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3290..3306
                     /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 gccaccatgc tgctatccgt
      121 gccgttgctg ctcggcctcc tcggcctggc cgtcgccctc gaggtgagcg agctgattaa
      181 ggagaacatg cacatgaagc tgtacatgga gggcaccgtg aacaaccacc acttcaagtg
      241 cacatccgag ggcgaaggca agccctacga gggcacccag accatgagaa tcaaggcggt
      301 cgagggcggc cctctcccct tcgccttcga catcctggct accagcttca tgtacggcag
      361 caaaaccttc atcaaccaca cccagggcat ccccgacttc tttaagcagt ccttccccga
      421 gggcttcaca tgggagagag tcaccacata cgaagacggg ggcgtgctga ccgctaccca
      481 ggacaccagc ctccaggacg gctgcctcat ctacaacgtc aagatcagag gggtgaactt
      541 cccatccaac ggccctgtga tgcagaagaa aacactcggc tgggaggcct ccaccgagac
      601 cctgtacccc gctgacggcg gcctggaagg cagagccgac atggccctga agctcgtggg
      661 cgggggccac ctgatctgca acttgaagac cacatacaga tccaagaaac ccgctaagaa
      721 cctcaagatg cccggcgtct actatgtgga cagaagactg gaaagaatca aggaggccga
      781 caaagagacc tacgtcgagc agcacgaggt ggctgtggcc agatactgcg acctccctag
      841 caaactgggg cacagaggat ccaaggacga gctgtaaacc cagctttctt gtacaaagtt
      901 ggcattataa gaaagcattg cttatcaatt tgttgcaacg aacaggtcac tatcagtcaa
      961 aataaaatca ttatttgcca tccagctgat atcccctata gtgagtcgta ttacatggtc
     1021 atagctgttt cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa
     1081 gataaaaata tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg
     1141 ggtgttatga gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat
     1201 gctgatttat atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc
     1261 tatcgcttgt atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc
     1321 gttgccaatg atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct
     1381 cttccgacca tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg
     1441 atccccggaa aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt
     1501 gttgatgcgc tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct
     1561 tttaacagcg atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg
     1621 gttgatgcga gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa
     1681 gaaatgcata aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca
     1741 cttgataacc ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc
     1801 ggaatcgcag accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct
     1861 ccttcattac agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa
     1921 ttgcagtttc atttgatgct cgatgagttt ttctaatcag aattggttaa ttggttgtaa
     1981 cactggcaga gcattacgct gacttgacgg gacggcgcaa gctcatgacc aaaatccctt
     2041 aacgtgagtt acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     2101 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     2161 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     2221 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     2281 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     2341 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     2401 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     2461 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     2521 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     2581 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     2641 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     2701 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     2761 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     2821 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
     2881 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     2941 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atacgcgtac cgctagccag
     3001 gaagagtttg tagaaacgca aaaaggccat ccgtcaggat ggccttctgc ttagtttgat
     3061 gcctggcagt ttatggcggg cgtcctgccc gccaccctcc gggccgttgc ttcacaacgt
     3121 tcaaatccgc tcccggcgga tttgtcctac tcaggagagc gttcaccgac aaacaacaga
     3181 taaaacgaaa ggcccagtct tccgactgag cctttcgttt tatttgatgc ctggcagttc
     3241 cctactctcg cgttaacgct agcatggatg ttttcccagt cacgacgttg taaaacgacg
     3301 gccagtctta agctcgggcc c
//
