LOCUS       sequence-446942-        3399 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A Gateway compatible middle clone with a Myc-tagged BIOID2 construct
            followed by a flexible linker and an MCS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3399)
  AUTHORS   Gillespie WB 3rd, Zhang Y, Ruiz OE, Cerda J 3rd, Ortiz-Guzman J, 
            Sherman M, Turner WD, Largoza G, Mosser LE, Fujimoto E, Chien CB, 
            Kwan KM, Arenkiel BR, Devine WP, Wythe JD
  TITLE     MultiSite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox for use in vertebrate model systems.
  JOURNAL   Development. 2025 Nov 15;152(22):dev204308. doi: 10.1242/dev.204308.
            Epub 2025 Nov 13.
  PUBMED    41230856
REFERENCE   2  (bases 1 to 3399)
  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: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: pME-Myc-BioID2-MCS2 (JDW 1209)
FEATURES             Location/Qualifiers
     source          1..3399
                     /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      693..702
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             702..731
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             735..1430
                     /codon_start=1
                     /product="promiscuous R40G mutant of a biotin protein 
                     ligase from Aquifex aeolicus (Kim et al., 2016)"
                     /label=BioID2
                     /translation="FKNLIWLKEVDSTQERLKEWNVSYGTALVADRQTKGRGGLGRKWL
                     SQEGGLYFSFLLNPKEFENLLQLPLVLGLSVSEALEEITEIPFSLKWPNDVYFQEKKVS
                     GVLCELSKDKLIVGIGINVNQREIPEEIKDRATTLYEITGKDWDRKEVLLKVLKRISEN
                     LKKFKEKSFKEFKGKIESKMLYLGEEVKLLGEGKITGKLVGLSEKGGALILTEEGIKEI
                     LSGEFSLRRS"
     protein_bind    complement(1524..1623)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1640..1659)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1641..1659)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1664..1680)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1664..1680)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1782..1801)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1793..2602
                     /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(1860..1879)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2749..3337
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3238..3257
                     /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 agcaggctac tagtgtcgac aagcttatcg atgccaccat ggaacaaaaa ctcatctcag
      721 aagaggatct cgacttcaag aacctgatct ggctgaagga ggtggacagc acccaggaga
      781 gactgaagga gtggaacgtg agctacggca ccgccctggt ggccgacaga cagaccaagg
      841 gcagaggcgg cctgggcaga aagtggctga gccaggaggg cggcctgtac ttcagcttcc
      901 tgctgaaccc caaggagttc gagaacctgc tgcagctgcc cctggtgctg ggcctgagcg
      961 tgagcgaggc cctggaggag atcaccgaga tccccttcag cctgaagtgg cccaacgacg
     1021 tgtacttcca ggagaagaag gtgagcggcg tgctgtgcga gctgagcaag gacaagctga
     1081 tcgtgggcat cggcatcaac gtgaaccaga gagagatccc cgaggagatc aaggacagag
     1141 ccaccaccct gtacgagatc accggcaagg actgggacag aaaggaggtg ctgctgaagg
     1201 tgctgaagag aatcagcgag aacctgaaga agttcaagga gaagagcttc aaggagttca
     1261 agggcaagat cgagagcaag atgctgtacc tgggcgagga ggtgaagctg ctgggcgagg
     1321 gcaagatcac cggcaagctg gtgggcctga gcgagaaggg cggcgccctg atcctgaccg
     1381 aggagggcat caaggagatc ctgagcggcg agttcagcct gagaagaagc accggtggat
     1441 caggaggatc ggcgcgccgt ggagcggccg ccgctagcta atgactcgag tgataaggta
     1501 cctgataaag atcttacgta tctagacagc tttcttgtac aaagttggca ttataagaaa
     1561 gcattgctta tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat
     1621 ttgccatcca gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg
     1681 gcagctctgg cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc
     1741 atcatgaaca ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca
     1801 tattcaacgg gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg
     1861 gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg
     1921 gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt
     1981 tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa
     2041 gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac
     2101 agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc
     2161 agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg
     2221 cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga
     2281 ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact
     2341 tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat
     2401 ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg
     2461 ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa
     2521 acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt
     2581 gatgctcgat gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat
     2641 tacgctgact tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc
     2701 gtcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
     2761 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
     2821 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
     2881 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
     2941 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
     3001 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3061 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
     3121 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
     3181 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
     3241 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
     3301 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
     3361 acggttcctg gccttttgct ggccttttgc tcacatgtt
//
