LOCUS       sequence-349871-        6066 bp    DNA     circular SYN 15-APR-2025
DEFINITION  encodes human NKD2 S31A mutant cDNA with a C-terminal mCherry 
            fusion..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6066)
  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 6066)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: pN1mcherry-NKD2S31A
FEATURES             Location/Qualifiers
     source          1..6066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        106..409
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        410..613
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     564..584
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1978..2004
                     /codon_start=1
                     /product="9xHis affinity tag"
                     /label=9xHis
                     /translation="HHHHHHHHH"
     regulatory      2062..2071
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2068..2778
                     /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(2213..2231)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2481..2500
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    2899..3020
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2936..2955)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2990..3009
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3027..3482)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3164..3185)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3376..3395
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3509..3613
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3615..3972
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3615..3806
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3638..3658)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3823..3958
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4007..4801
                     /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             4007..4801
                     /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(4061..4080)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4671..4690
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4989..5008)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5033..5080
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5409..5997
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5898..5917
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcgttatccc ctgattctgt ggataaccgt attaccgcca tgcattagtt attaatagta
       61 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
      121 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
      181 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
      241 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
      301 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
      361 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
      421 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
      481 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
      541 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
      601 tataagcaga gctggtttag tgaaccgtca gatccgctag cgctaccgga ctcagatctc
      661 gagatgggga aactgcagtc gaagcacgcc gccgccgccc gcaagcggag agagagcccg
      721 gaaggggaca gcttcgtggc gtccgcgtac gcggccggcc gcaaaggcgc ggaggaagcg
      781 gagcggcgcg cgcgggacaa gcaggagctg cccaatgggg accccaagga ggggcctttc
      841 cgggaggacc agtgtcccct acaggtggca ctccccgctg agaaagctga gggccgcgag
      901 cacccgggac aactcctcag cgcagatgac ggagagaggg cagcaaaccg cgagggcccg
      961 cgaggaccgg gcgggcagcg cctcaacatt gacgcactcc agtgcgatgt ctcggtggag
     1021 gaggacgacc gccaggagtg gacgttcacg ctctatgact ttgacaactg cgggaaggtc
     1081 accagggagg acatgtccag cctcatgcac accatctatg aggtcgtgga tgcctcggtc
     1141 aaccactcct cgggcagcag caagaccctc cgtgtgaagc taaccgtcag ccctgagccc
     1201 tccagcaaga ggaaggaggg tcctcctgct ggccaggacc gggagcccac ccgttgcagg
     1261 atggagggtg aactggcaga ggagccaagg gtggctgaca ggaggttgtc tgcacacgtc
     1321 aggaggccca gtactgaccc ccagccctgc tcggagcggg ggccctactg cgtggacgag
     1381 aacacggagc gcagaaacca ctacctggac ctcgccggga ttgagaacta cacgtccaga
     1441 ttcggccctg ggtcccctcc tgtgcaagca aagcaggagc cccagggcag ggcctcgcac
     1501 ctccaggccc ggtcccgctc ccaggagcca gatacacatg ccgtacacca ccgcaggtca
     1561 caggtgctgg tggaacacgt cgtgccagcc tcggagcctg ctgcccgggc cctggacacg
     1621 cagccccggc cgaaggggcc ggagaagcag ttcctcaagt cccccaaggg ctccgggaag
     1681 ccgcctgggg tgccagccag cagcaagtcc gggaaagcct tcagctacta cctgccggcc
     1741 gtcctgccgc cccaggcccc tcaggacggc caccacctcc cgcagccccc accgccaccc
     1801 tacggccaca agcggtaccg ccaaaagggc agggagggcc actcgccact caaggcccca
     1861 cacgctcagc ctgccacagt ggagcacgag gtggtgcggg acctgccgcc cacgccagca
     1921 ggagagggct acgcggtgcc agtgatccag cggcacgagc accaccacca ccacgagcac
     1981 caccaccacc accaccacca ccacttccac ccgtcccgaa ttctgcagtc gacggtaccg
     2041 cgggcccggg atccaccggt cgccaccatg gtgagcaagg gcgaggagga taacatggcc
     2101 atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag
     2161 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg
     2221 aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg
     2281 tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc
     2341 ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc
     2401 gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc
     2461 accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc
     2521 tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag
     2581 ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc
     2641 gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag
     2701 gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg
     2761 gacgagctgt acaagtagcg gccgcgactc tagatcataa tcagccatac cacatttgta
     2821 gaggttttac ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg
     2881 aatgcaattg ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     2941 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     3001 aaactcatca atgtatctta aggcgtaaat tgtaagcgtt aatattttgt taaaattcgc
     3061 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     3121 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     3181 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     3241 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     3301 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     3361 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     3421 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     3481 gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     3541 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     3601 aaaaaggaag agtcctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt
     3661 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3721 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3781 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3841 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3901 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3961 ggcttttgca aagatcgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     4021 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     4081 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     4141 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcaag acgaggcagc
     4201 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     4261 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     4321 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     4381 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     4441 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     4501 cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg cccgacggcg aggatctcgt
     4561 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     4621 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     4681 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     4741 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     4801 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     4861 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     4921 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccctaggggg
     4981 aggctaactg aaacacggaa ggagacaata ccggaaggaa cccgcgctat gacggcaata
     5041 aaaagacaga ataaaacgca cggtgttggg tcgtttgttc ataaacgcgg ggttcggtcc
     5101 cagggctggc actctgtcga taccccaccg agaccccatt ggggccaata cgcccgcgtt
     5161 tcttcctttt ccccacccca ccccccaagt tcgggtgaag gcccagggct cgcagccaac
     5221 gtcggggcgg caggccctgc catagcctca ggttactcat atatacttta gattgattta
     5281 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     5341 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     5401 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     5461 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     5521 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
     5581 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     5641 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     5701 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     5761 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     5821 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     5881 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     5941 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     6001 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     6061 tttcct
//
