LOCUS       sequence-79771-e         926 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses human NKCC1 V740C mutant with an N-terminal 3xFLAG-YFP tag
            in mammalian cells. cDNA is synthetic, containing convenient 
            restriction sites..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 926)
  TITLE     Forbush Lab human NKCC1 NT clones
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 926)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pcDNA3.1 Flag YFP hNKCC1 V740C (NT526)
FEATURES             Location/Qualifiers
     source          1..926
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     regulatory      6..15
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             18..83
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             join(96..98,99..809)
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(96..98,99..809)
                     /codon_start=1
                     /product="enhanced YFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(138..159)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(399..418)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     746..767
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 cggccgccac catgggagac tacaaagacc atgacggtga ttataaagat cacgacatcg
       61 actacaagga tgacgatgac aagggtgacc ctaccgtcag caagggcgag gagctgttca
      121 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
      181 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
      241 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccttcggc tacggcctga
      301 tgtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
      361 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
      421 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
      481 acttcaagga ggacggcaac atcctagggc acaagctgga gtacaactac aacagccaca
      541 acgtttatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
      601 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
      661 gcgacggccc cgtgctgctg cccgacaacc actacctgag ctaccagtcc aaactgagca
      721 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
      781 tcactctcgg catggacgag ctctacaagg gtgacctgga gccgaggccc acggctccta
      841 gctccggcgc gccaggcctc gccggagtcg gcgaaacgcc gagcgccgct gcgctggccg
      901 cagccagggt ggaactgccc ggaacc
//
