LOCUS       sequence-437200-        3267 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway entry clone with codon-optimized ORF sequence for subcloning
            into custom expression vectors.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3267)
  AUTHORS   Superti-Furga G, Lackner D, Wiedmer T, Ingles-Prieto A, Barbosa B, 
            Girardi E, Goldmann U, Gurtl B, Klavins K, Klimek C, Lindinger S, 
            Lineiro-Retes E, Muller AC, Onstein S, Redinger G, Reil D, Sedlyarov
            V, Wolf G, Crawford M, Everley R, ... Steppan CM
  TITLE     The RESOLUTE consortium: unlocking SLC transporters for drug 
            discovery.
  JOURNAL   Nat Rev Drug Discov. 2020 Jul;19(7):429-430. doi: 
            10.1038/d41573-020-00056-6.
  PUBMED    32265506
REFERENCE   2  (bases 1 to 3267)
  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"; doi: "10"; journalName: 
            "Nat Rev Drug Discov"; date: "2020-07"; volume: "19"; issue: "7"; 
            pages: "429-430"
COMMENT     Sequence Label: pDONR221-eGFP
FEATURES             Location/Qualifiers
     source          1..3267
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     94..111
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(270..297)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(389..475)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     524..546
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     538..555
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     539..555
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    571..670
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(676..678,679..681,682..1392)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(721..742)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(982..1001)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1329..1350
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1394..1493)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1510..1529)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1511..1529)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1534..1550)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1534..1550)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1652..1671)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1663..2472
                     /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(1730..1749)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2619..3207
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3108..3127
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      181 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tacgcgtacc
      241 gctagccagg aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg gccttctgct
      301 tagtttgatg cctggcagtt tatggcgggc gtcctgcccg ccaccctccg ggccgttgct
      361 tcacaacgtt caaatccgct cccggcggat ttgtcctact caggagagcg ttcaccgaca
      421 aacaacagat aaaacgaaag gcccagtctt ccgactgagc ctttcgtttt atttgatgcc
      481 tggcagttcc ctactctcgc gttaacgcta gcatggatgt tttcccagtc acgacgttgt
      541 aaaacgacgg ccagtcttaa gctcgggccc caaataatga ttttattttg actgatagtg
      601 acctgttcgt tgcaacaaat tgatgagcaa tgctttttta taatgccaac tttgtacaaa
      661 aaagcaggct ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
      721 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
      781 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
      841 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
      901 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
      961 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     1021 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     1081 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     1141 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     1201 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     1261 ctgcccgaca accactacct gagcacccag tccaagctga gcaaagaccc caacgagaag
     1321 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac
     1381 gagctgtaca aggacccagc tttcttgtac aaagttggca ttataagaaa gcattgctta
     1441 tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat ttgccatcca
     1501 gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg
     1561 cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgaaca
     1621 ataaaactgt ctgcttacat aaacagtaat acaaggggtg ttatgagcca tattcaacgg
     1681 gaaacgtcga ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg
     1741 gctcgcgata atgtcgggca atcaggtgcg acaatctatc gcttgtatgg gaagcccgat
     1801 gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag
     1861 atggtcagac taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc
     1921 cgtactcctg atgatgcatg gttactcacc actgcgatcc ccggaaaaac agcattccag
     1981 gtattagaag aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg
     2041 cgccggttgc attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt
     2101 ctcgctcagg cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac
     2161 gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa tgcataaact tttgccattc
     2221 tcaccggatt cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag
     2281 gggaaattaa taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat
     2341 cttgccatcc tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt
     2401 caaaaatatg gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat
     2461 gagtttttct aatcagaatt ggttaattgg ttgtaacact ggcagagcat tacgctgact
     2521 tgacgggacg gcgcaagctc atgaccaaaa tcccttaacg tgagttacgc gtcgttccac
     2581 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2641 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2701 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2761 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2821 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2881 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2941 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3001 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     3061 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     3121 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     3181 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     3241 gccttttgct ggccttttgc tcacatg
//
