LOCUS       sequence-342344-        4327 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Multisite Gateway middle entry clone encoding TurboID with mKate2 
            marker. Parton lab clone IYN.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4327)
  AUTHORS   Xiong Z, Lo HP, McMahon KA, Martel N, Jones A, Hill MM, Parton RG, 
            Hall TE
  TITLE     In vivo proteomic mapping through GFP-directed proximity-dependent 
            biotin labelling in zebrafish.
  JOURNAL   Elife. 2021 Feb 16;10. pii: 64631. doi: 10.7554/eLife.64631.
  PUBMED    33591275
REFERENCE   2  (bases 1 to 4327)
  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: "Elife."; 
            date: "2021-02-16"; volume: "10. pii: 64631. doi"; pages: " 
            10.7554/eLife.64631"
COMMENT     Sequence Label: pME-mKate2-P2A-TurboID-NS
FEATURES             Location/Qualifiers
     source          1..4327
                     /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    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      669..678
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             675..1370
                     /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"
     CDS             1386..1442
                     /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             1461..1490
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    complement(2452..2551)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2568..2587)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2569..2587)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2592..2608)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2592..2608)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2710..2729)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2721..3530
                     /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(2788..2807)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3677..4265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4166..4185
                     /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 tgtacaaaaa
      661 agcaggctgc caccatggtg agcgagctga ttaaggagaa catgcacatg aagctgtaca
      721 tggagggcac cgtgaacaac caccacttca agtgcacatc cgagggcgaa ggcaagccct
      781 acgagggcac ccagaccatg agaatcaagg cggtcgaggg cggccctctc cccttcgcct
      841 tcgacatcct ggctaccagc ttcatgtacg gcagcaaaac cttcatcaac cacacccagg
      901 gcatccccga cttctttaag cagtccttcc ccgagggctt cacatgggag agagtcacca
      961 catacgaaga cgggggcgtg ctgaccgcta cccaggacac cagcctccag gacggctgcc
     1021 tcatctacaa cgtcaagatc agaggggtga acttcccatc caacggccct gtgatgcaga
     1081 agaaaacact cggctgggag gcctccaccg agaccctgta ccccgctgac ggcggcctgg
     1141 aaggcagagc cgacatggcc ctgaagctcg tgggcggggg ccacctgatc tgcaacttga
     1201 agaccacata cagatccaag aaacccgcta agaacctcaa gatgcccggc gtctactatg
     1261 tggacagaag actggaaaga atcaaggagg ccgacaaaga gacctacgtc gagcagcacg
     1321 aggtggctgt ggccagatac tgcgacctcc ctagcaaact ggggcacaga cctaggggca
     1381 gcggcgccac caacttcagc ctgctgaagc aggccggcga cgtggaggag aaccccggcc
     1441 ccgcccccgg cagcctcgag gagcagaaac tgataagcga agaggacctg aaagacaata
     1501 cagtcccctt gaagctgatt gcccttctgg caaatggcga attccacagc ggggagcaac
     1561 tgggcgaaac gttgggcatg agtagagcag cgataaacaa acatatacaa acgctgaggg
     1621 attggggtgt agacgtgttc accgttccgg gcaagggcta tagtttgccc gaaccgatac
     1681 cacttctgaa cgccaaacaa atattgggtc aactggatgg aggatcagta gcagttcttc
     1741 ctgtggtcga ctctactaac caatatctgc tggatcggat cggggaactg aaatctggcg
     1801 atgcatgcat agctgagtac cagcaggctg gacggggatc tcgaggaagg aaatggtttt
     1861 ccccgtttgg ggccaacctc tatctctcta tgttctggag gttgaaacgt ggccctgctg
     1921 ccatcggctt ggggcctgtc attggtattg ttatggcgga ggcgttgcgt aaacttggag
     1981 cggataaagt gagggttaaa tggccaaacg atttgtatct tcaagaccgc aagttggctg
     2041 gtatactggt agaactcgca gggattacag gagatgcggc tcagattgtg atcggtgccg
     2101 gtattaacgt cgcgatgcgt agggtagagg agtccgtggt caatcagggt tggattacgc
     2161 tgcaagaggc tggaatcaat ctcgacagaa acacacttgc agccacgctt attcgggaat
     2221 tgagggctgc tcttgaactt tttgaacagg aagggttggc accttacctg ccgcgctggg
     2281 aaaagctgga taatttcata aatcggcccg tcaagctcat aatcggggac aaggaaattt
     2341 tcggcatatc acgcggcatc gacaagcagg gggcactgct gttggaacag gacggagtca
     2401 taaagccctg gatgggtggt gagattagcc ttcgatctgc ggagaaagtc gacccagctt
     2461 tcttgtacaa agttggcatt ataagaaagc attgcttatc aatttgttgc aacgaacagg
     2521 tcactatcag tcaaaataaa atcattattt gccatccagc tgatatcccc tatagtgagt
     2581 cgtattacat ggtcatagct gtttcctggc agctctggcc cgtgtctcaa aatctctgat
     2641 gttacattgc acaagataaa aatatatcat catgaacaat aaaactgtct gcttacataa
     2701 acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcgagg ccgcgattaa
     2761 attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
     2821 caggtgcgac aatctatcgc ttgtatggga agcccgatgc gccagagttg tttctgaaac
     2881 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga
     2941 cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt
     3001 tactcaccac tgcgatcccc ggaaaaacag cattccaggt attagaagaa tatcctgatt
     3061 caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg
     3121 tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa
     3181 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
     3241 aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc
     3301 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg
     3361 atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc
     3421 tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc
     3481 ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa tcagaattgg
     3541 ttaattggtt gtaacactgg cagagcatta cgctgacttg acgggacggc gcaagctcat
     3601 gaccaaaatc ccttaacgtg agttacgcgt cgttccactg agcgtcagac cccgtagaaa
     3661 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     3721 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     3781 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     3841 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     3901 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     3961 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     4021 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     4081 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     4141 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     4201 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     4261 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     4321 acatgtt
//
