LOCUS       sequence-178084-        7424 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MultiSite Gateway entry clones, second fragment (attL5, attL2) Human
            NFAT5 CDS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7424)
  AUTHORS   Jackel C, Nogueira MS, Ehni N, Kraus C, Ranke J, Dohmann M, Noessner
            E, Nelson PJ
  TITLE     A vector platform for the rapid and efficient engineering of stable 
            complex transgenes.
  JOURNAL   Sci Rep. 2016 Oct 3;6:34365. doi: 10.1038/srep34365.
  PUBMED    27694838
REFERENCE   2  (bases 1 to 7424)
  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: "Sci Rep."; 
            date: "2016-10-3"; volume: "6:34365. doi"; pages: " 
            10.1038/srep34365"
COMMENT     Sequence Label: pENTR221-NFAT5
FEATURES             Location/Qualifiers
     source          1..7424
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    570..665
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        680..698
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      733..742
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             742..765
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    complement(5536..5635)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5665..5684)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5666..5684)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5689..5705)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5689..5705)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5807..5826)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5818..6627
                     /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(5885..5904)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      6774..7362
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7263..7282
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtatacaaa
      661 agttgtacgc caagctcgaa attaaccctc actaaaggga acaaaagctg gagctccacc
      721 gcggtggcgg ccgccaccat ggattacaag gatgacgacg ataagaagcc cgggcggatc
      781 ctgccctcgg acttcatctc attgctcagc gcggacctag acctggaatc gcccaagtcc
      841 ctctactcgc gagaatctgt ctatgatctt ctcccaaagg agttacagtt acctccatct
      901 agagaaacat ctgtagcatc aatgagtcag acaagcggtg gtgaggcagg ctcgcctcct
      961 ccagctgttg ttgctgctga tgcttcttca gctccctcct cttcctccat gggcggtgct
     1021 tgcagctcct ttaccacctc ttccagccct accatttatt ctacctcagt caccgacagc
     1081 aaggctatgc aagtggagag ctgctcctca gccgtggggg taagtaacag aggggtaagt
     1141 gaaaagcagt taaccagtaa cacagttcag cagcatccat caacaccgaa gaggcacaca
     1201 gtcttgtaca tctcaccacc acctgaggac ttgctggata acagtcggat gtcctgccag
     1261 gatgaggggt gtggattgga atctgagcag agctgcagta tgtggatgga ggattccccc
     1321 tccaacttca gtaacatgag caccagttcc tacaatgata acactgaggt acctcgtaaa
     1381 tcacgaaaac gaaatccaaa gcagaggccg ggggtcaaac gacgagattg tgaagaatct
     1441 aatatggata tatttgatgc cgacagtgcc aaagcacctc actatgtgct ttctcagctt
     1501 accacggaca acaaaggcaa ctcaaaagcg ggaaatggaa cattggaaaa ccaaaaagga
     1561 actggagtaa agaagagccc tatgttgtgt ggacaatatc ctgttaaaag tgagggaaag
     1621 gagctgaaga tagttgtaca acctgagaca cagcaccgag ctcggtacct gactgagggc
     1681 agccgtggct cagtgaaaga tagaacacag caaggctttc ctacagtaaa gctggaaggc
     1741 cataatgaac ctgtagtgtt gcaagtgttt gtgggcaacg actctggacg agtgaaacca
     1801 catggatttt atcaggcctg cagagtaact ggacgaaata caactccttg caaagaagtg
     1861 gacattgaag gcactactgt tatagaagtc ggccttgatc ctagcaacaa catgacactg
     1921 gcggtggact gcgtagggat attgaaattg aggaatgctg atgtcgaagc cagaatagga
     1981 attgctggtt ccaagaagaa aagcactcgt gccagattgg tttttcgagt taatatcatg
     2041 aggaaagatg gctccacttt gacactgcaa acaccctctt ctccaatttt gtgtactcag
     2101 ccagcaggag tgccagaaat cttaaagaaa agcttgcata gctgttcagt gaaaggagaa
     2161 gaagaagtgt ttttaatcgg caagaacttt ctgaaaggaa ctaaagttat tttccaagaa
     2221 aatgtttctg atgaaaactc ttggaagtca gaagctgaaa ttgatatgga actatttcat
     2281 cagaatcatc ttattgtgaa ggttcctccc tatcatgacc aacatataac tttgcctgtg
     2341 tcagtgggaa tatatgtagt gacaaatgct ggaagatctc atgatgttca accattcact
     2401 tacactccag acccagcagc agctggtgct ttgaatgtaa atgtgaagaa ggaaatatct
     2461 agtccagcaa gaccttgctc ttttgaagag gccatgaaag caatgaaaac tactggatgt
     2521 aatttagata aggtaaatat tatccctaat gccctgatga ctccactcat accaagcagt
     2581 atgattaaga gtgaagatgt tactccaatg gaagtaacag cagaaaaaag atcttccact
     2641 atttttaaga ctacaaagtc tgttggatca actcagcaaa cattagaaaa catctcaaac
     2701 atagcaggaa atggctcttt ttcatcacca tcatcttccc acctaccttc tgaaaatgaa
     2761 aaacagcagc agattcagcc caaggcatac aacccagaga ccctgacaac tattcaaacc
     2821 caggacatct cacagcctgg tacttttcca gcagtttctg cttctagtca gctgcccaac
     2881 agcgatgcac tattgcagca ggctacacag tttcagacaa gagaaactca gtctagagag
     2941 atattacagt cagatggtac agtggttaat ttgtcacaac tgactgaggc atcacaacaa
     3001 cagcagcagt caccactaca agaacaagca cagactttac agcagcagat ttcatcaaat
     3061 atttttccat caccaaatag tgtgagtcag cttcagaata ctattcagca gctgcaagca
     3121 gggagtttca caggcagtac tgctagtggc agcagtggaa gtgttgactt ggtccaacaa
     3181 gttttagagg cacagcagca gttatcttca gttttatttt ctgctccaga tggtaatgag
     3241 aatgttcaag agcagcttag tgcagatatt tttcaacaag tcagtcaaat tcagagtggt
     3301 gtaagccctg gaatgttttc ctcaacagag ccaacagtcc ataccagacc agataattta
     3361 ttacctggaa gagctgaaag tgttcatcca cagtctgaaa acacgttatc taatcaacag
     3421 cagcagcagc agcagcaaca gcaagtgatg gaatcttcag ccgcaatggt gatggagatg
     3481 caacagagta tctgccaggc agctgcccag attcagtcag agttattccc ttcaactgct
     3541 tcagcaaatg gaaaccttca gcaatcgcca gtttaccagc agacttctca catgatgagt
     3601 gcattgtcta ccaatgagga tatgcaaatg cagtgtgaat tgttttcttc tcctcctgca
     3661 gtttctggaa atgaaacttc tacaactacc acacagcagg ttgcaacccc tggcactacc
     3721 atgtttcaga catcaagttc aggagatgga gaagaaactg gaacacaagc aaaacagatt
     3781 cagaacagtg tctttcagac catggtccaa atgcaacata gtggggacaa tcaacctcaa
     3841 gttaaccttt tttcatccac aaaaagtatg atgagtgttc agaatagtgg tacccaacaa
     3901 caaggtaatg gtttattcca gcaagggaat gagatgatgt cacttcaatc tggaaatttt
     3961 ttgcagcagt cttctcattc acaggcccaa ctttttcatc ctcaaaatcc tattgccgat
     4021 gctcagaacc tttcccagga aactcaaggt tctctctttc atagtccaaa tcctattgtc
     4081 cacagtcaga cttctacaac ctcctctgaa caaatgcagc ctccaatgtt tcactctcaa
     4141 agtaccattg ctgtgttaca gggctcttca gttcctcaag accagcagtc aaccaacata
     4201 tttctttccc agagtcccat gaataatctt cagactaaca cagtagccca agaagcattt
     4261 tttgcagcac cgaactcaat ttctccactt cagtcaacat caaacagtga acaacaagct
     4321 gctttccaac agcaagctcc aatatcacac atccagactc ctatgctttc ccaagaacag
     4381 gcacaacccc cgcagcaggg tttatttcag cctcaggtgg ccctgggctc ccttccacct
     4441 aatccaatgc ctcaaagcca acaaggaacc atgttccagt cacagcactc aatagttgcc
     4501 atgcagagta actctccatc ccaggaacag cagcagcagc agcaacagca gcagcaacag
     4561 cagcagcaac aacaacagag cattttattc agtaatcaga ataccatggc tacaatggcg
     4621 tctccaaagc aaccaccacc aaacatgata ttcaacccaa atcaaaatcc aatggctaat
     4681 caggagcaac agaaccagtc aatttttcac caacaaagta acatggcccc aatgaatcaa
     4741 gagcaacagc ccatgcaatt tcagagtcag tccacagttt cctcacttca gaacccaggt
     4801 cctacccagt cggaatcatc acagaccccc ttgttccata gctctcctca gattcagttg
     4861 gtacaagggt cacctagttc tcaagagcag caagtaactc tcttcttatc tccagcatcc
     4921 atgtctgcct tgcagaccag tataaatcaa caagatatgc aacagtctcc tctttattcc
     4981 cctcagaaca acatgcctgg aattcaagga gccacatctt cgcctcaacc acaggctact
     5041 ttatttcaca acacagcagg aggcacaatg aaccaactgc agaattctcc tggctcatct
     5101 cagcagacat caggaatgtt cttatttggc attcaaaata actgtagtca gcttttaacc
     5161 tctggaccag ctacattgcc tgatcagttg atggccataa gtcagccagg ccaaccacaa
     5221 aacgagggcc agccacctgt gacaacactt ctttctcagc aaatgccaga gaattctcca
     5281 ctggcatcct ctataaacac caaccagaac atcgaaaaga ttgatttgct tgtttcattg
     5341 caaaaccaag ggaacaactt gactggctcc ttttaactgg atataaattc cacgaagaaa
     5401 atcctgattc caagatgtcc tgagatcttg tggttccatg agaattatta ctttaaaaaa
     5461 aaagtcgacc tcgagggggg gcccggtacc ttaattaatt aaggtaccag gtaagtgtac
     5521 ccaattcgcc ctataaccca gctttcttgt acaaagttgg cattataaga aagcattgct
     5581 tatcaatttg ttgcaacgaa caggtcacta tcagtcaaaa taaaatcatt atttgccatc
     5641 cagctgcagg gcggccgcga tatcccctat agtgagtcgt attacatggt catagctgtt
     5701 tcctggcagc tctggcccgt gtctcaaaat ctctgatgtt acattgcaca agataaaaat
     5761 atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     5821 agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga tgctgattta
     5881 tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat ctatcgcttg
     5941 tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat
     6001 gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc tcttccgacc
     6061 atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccgga
     6121 aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg
     6181 ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc ttttaacagc
     6241 gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg
     6301 agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat
     6361 aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac
     6421 cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca
     6481 gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc tccttcatta
     6541 cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt
     6601 catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta acactggcag
     6661 agcattacgc tgacttgacg ggacggcgca agctcatgac caaaatccct taacgtgagt
     6721 tacgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     6781 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     6841 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     6901 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     6961 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     7021 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     7081 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     7141 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     7201 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     7261 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     7321 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     7381 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgtt
//
