LOCUS       sequence-486041-        3308 bp    DNA     circular SYN 16-SEP-2025
DEFINITION  Middle entry fluorescent mCherry reporter with a viral 2A peptide 
            for use with the Gateway cloning system; allows 3' transgene to be 
            transcribed on the same mRNA but translated separately.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3308)
  AUTHORS   Kendall GC, Watson S, Xu L, LaVigne CA, Murchison W, Rakheja D, 
            Skapek SX, Tirode F, Delattre O, Amatruda JF
  TITLE     PAX3-FOXO1 transgenic zebrafish models identify HES3 as a mediator 
            of rhabdomyosarcoma tumorigenesis.
  JOURNAL   eLife. 2018 Jun 5;7. pii: 33800. doi: 10.7554/eLife.33800.
  PUBMED    29869612
REFERENCE   2  (bases 1 to 3308)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 16, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "eLife."; 
            date: "2018-06-5"; volume: "7. pii: 33800. doi"; pages: " 
            10.7554/eLife.33800"
COMMENT     Sequence Label: mE-mCherry2A
FEATURES             Location/Qualifiers
     source          1..3308
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             19..726
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(164..182)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     432..451
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             727..780
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     protein_bind    complement(782..881)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(898..917)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(899..917)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(922..938)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(922..938)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1040..1059)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1051..1860
                     /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(1118..1137)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2007..2595
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2496..2515
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2749..2766
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2925..2952)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3044..3130)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3179..3201
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3193..3210
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3194..3210
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    join(3226..3308,1..17)
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
ORIGIN
        1 gtacaaaaaa gcaggcttat ggtgagcaag ggcgaggagg ataacatggc catcatcaag
       61 gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc
      121 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
      181 aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc
      241 aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc cttccccgag
      301 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag
      361 gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg caccaacttc
      421 ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc ctccgagcgg
      481 atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac
      541 ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg
      601 cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc
      661 atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg
      721 tacaaggagg gcagaggaag tcttctaaca tgcggtgacg tggaggagaa tcccggccct
      781 tacccagctt tcttgtacaa agttggcatt ataagaaagc attgcttatc aatttgttgc
      841 aacgaacagg tcactatcag tcaaaataaa atcattattt gccatccagc tgatatcccc
      901 tatagtgagt cgtattacat ggtcatagct gtttcctggc agctctggcc cgtgtctcaa
      961 aatctctgat gttacattgc acaagataaa aatatatcat catgaacaat aaaactgtct
     1021 gcttacataa acagtaatac aaggggtgtt atgagccata ttcaacggga aacgtcgagg
     1081 ccgcgattaa attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat
     1141 gtcgggcaat caggtgcgac aatctatcgc ttgtatggga agcccgatgc gccagagttg
     1201 tttctgaaac atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta
     1261 aactggctga cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat
     1321 gatgcatggt tactcaccac tgcgatcccc ggaaaaacag cattccaggt attagaagaa
     1381 tatcctgatt caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat
     1441 tcgattcctg tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg
     1501 caatcacgaa tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc
     1561 tggcctgttg aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca
     1621 gtcgtcactc atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata
     1681 ggttgtattg atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta
     1741 tggaactgcc tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt
     1801 attgataatc ctgatatgaa taaattgcag tttcatttga tgctcgatga gtttttctaa
     1861 tcagaattgg ttaattggtt gtaacactgg cagagcatta cgctgacttg acgggacggc
     1921 gcaagctcat gaccaaaatc ccttaacgtg agttacgcgt cgttccactg agcgtcagac
     1981 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2041 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2101 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     2161 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     2221 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     2281 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     2341 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     2401 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     2461 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     2521 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     2581 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     2641 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     2701 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     2761 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     2821 cattaatgca gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca
     2881 attaatacgc gtaccgctag ccaggaagag tttgtagaaa cgcaaaaagg ccatccgtca
     2941 ggatggcctt ctgcttagtt tgatgcctgg cagtttatgg cgggcgtcct gcccgccacc
     3001 ctccgggccg ttgcttcaca acgttcaaat ccgctcccgg cggatttgtc ctactcagga
     3061 gagcgttcac cgacaaacaa cagataaaac gaaaggccca gtcttccgac tgagcctttc
     3121 gttttatttg atgcctggca gttccctact ctcgcgttaa cgctagcatg gatgttttcc
     3181 cagtcacgac gttgtaaaac gacggccagt cttaagctcg ggccccaaat aatgatttta
     3241 ttttgactga tagtgacctg ttcgttgcaa caaattgatg agcaatgctt ttttataatg
     3301 ccaacttt
//
