LOCUS       sequence-349816-        6077 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  encodes human NKD2 cDNA with a C-terminal GFP-fusion..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6077)
  AUTHORS   Wu B, Woo JS, Vila P, Jew M, Leung J, Sun Z, Srikanth S, Gwack Y
  TITLE     NKD2 mediates stimulation-dependent ORAI1 trafficking to augment 
            Ca(2+) entry in T cells.
  JOURNAL   Cell Rep. 2021 Aug 24;36(8):109603. doi: 
            10.1016/j.celrep.2021.109603.
  PUBMED    34433025
REFERENCE   2  (bases 1 to 6077)
  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: "Cell Rep.";
            date: "2021-08-24"; volume: "36(8):109603. doi"; pages: " 
            10.1016/j.celrep.2021.109603"
COMMENT     Sequence Label: pEGFPN1-hNKD2
FEATURES             Location/Qualifiers
     source          1..6077
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        94..397
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        398..601
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     552..572
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1966..1992
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     regulatory      2050..2059
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2056..2058,2059..2061,2062..2775)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2056..2058,2059..2061,2062..2772)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2101..2122)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2362..2381)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2709..2730
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2898..3019
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2935..2954)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2989..3008
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3026..3481)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3163..3184)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3375..3394
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3508..3612
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3614..3971
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3614..3805
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3637..3657)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3822..3957
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4006..4800
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4006..4800
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4060..4079)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4670..4689
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4988..5007)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5032..5079
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5408..5996
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5897..5916
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat caattacggg
       61 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      121 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      181 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      241 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
      301 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
      361 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
      421 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      481 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
      541 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      601 tggtttagtg aaccgtcaga tccgctagcg ctaccggact cagatctcga gatggggaaa
      661 ctgcagtcga agcacgccgc cgccgcccgc aagcggagag agagcccgga aggggacagc
      721 ttcgtggcgt ccgcgtacgc gagcggccgc aaaggcgcgg aggaagcgga gcggcgcgcg
      781 cgggacaagc aggagctgcc caatggggac cccaaggagg ggcctttccg ggaggaccag
      841 tgtcccctac aggtggcact ccccgctgag aaagctgagg gccgcgagca cccgggacaa
      901 ctcctcagcg cagatgacgg agagagggca gcaaaccgcg agggcccgcg aggaccgggc
      961 gggcagcgcc tcaacattga cgcactccag tgcgatgtct cggtggagga ggacgaccgc
     1021 caggagtgga cgttcacgct ctatgacttt gacaactgcg ggaaggtcac cagggaggac
     1081 atgtccagcc tcatgcacac catctatgag gtcgtggatg cctcggtcaa ccactcctcg
     1141 ggcagcagca agaccctccg tgtgaagcta accgtcagcc ctgagccctc cagcaagagg
     1201 aaggagggtc ctcctgctgg ccaggaccgg gagcccaccc gttgcaggat ggagggtgaa
     1261 ctggcagagg agccaagggt ggctgacagg aggttgtctg cacacgtcag gaggcccagt
     1321 actgaccccc agccctgctc ggagcggggg ccctactgcg tggacgagaa cacggagcgc
     1381 agaaaccact acctggacct cgccgggatt gagaactaca cgtccagatt cggccctggg
     1441 tcccctcctg tgcaagcaaa gcaggagccc cagggcaggg cctcgcacct ccaggcccgg
     1501 tcccgctccc aggagccaga tacacatgcc gtacaccacc gcaggtcaca ggtgctggtg
     1561 gaacacgtcg tgccagcctc ggagcctgct gcccgggccc tggacacgca gccccggccg
     1621 aaggggccgg agaagcagtt cctcaagtcc cccaagggct ccgggaagcc gcctggggtg
     1681 ccagccagca gcaagtccgg gaaagccttc agctactacc tgccggccgt cctgccgccc
     1741 caggcccctc aggacggcca ccacctcccg cagcccccac cgccacccta cggccacaag
     1801 cggtaccgcc aaaagggcag ggagggccac tcgccactca aggccccaca cgctcagcct
     1861 gccacagtgg agcacgaggt ggtgcgggac ctgccgccca cgccagcagg agagggctac
     1921 gcggtgccag tgatccagcg gcacgagcac caccaccacc acgagcacca ccaccaccac
     1981 caccaccacc acttccaccc gtcccgaatt ctgcagtcga cggtaccgcg ggcccgggat
     2041 ccaccggtcg ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc
     2101 ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag
     2161 ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc
     2221 gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac
     2281 cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag
     2341 gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc
     2401 gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc
     2461 aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc
     2521 gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc
     2581 agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg
     2641 ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag
     2701 cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac
     2761 gagctgtaca agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag
     2821 aggttttact tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga
     2881 atgcaattgt tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     2941 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     3001 aactcatcaa tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg
     3061 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     3121 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     3181 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     3241 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     3301 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     3361 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     3421 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     3481 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     3541 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     3601 aaaaggaaga gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg
     3661 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     3721 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3781 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     3841 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     3901 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     3961 gcttttgcaa agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat
     4021 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
     4081 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
     4141 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg
     4201 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
     4261 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
     4321 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
     4381 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
     4441 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
     4501 gcgccagccg aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc
     4561 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
     4621 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
     4681 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
     4741 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
     4801 gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt
     4861 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     4921 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga
     4981 ggctaactga aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa
     5041 aaagacagaa taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc
     5101 agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt
     5161 cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg
     5221 tcggggcggc aggccctgcc atagcctcag gttactcata tatactttag attgatttaa
     5281 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     5341 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     5401 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     5461 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     5521 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     5581 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     5641 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     5701 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     5761 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     5821 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     5881 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     5941 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     6001 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     6061 ttcctgcgtt atcccct
//
