LOCUS       sequence-217104-        5922 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For mammalian expression of the human NODAL open reading frame (with
            two mutated N-glycosylation sites) with a C-terminal MYC-DYK tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5922)
  AUTHORS   Findlay SD, Bilyk O, Lypka K, Waskiewicz AJ, Postovit L
  TITLE     Genetically regulated human NODAL splice variants are differentially
            post-transcriptionally processed and functionally distinct
  JOURNAL   (2018) bioRxiv 276170
REFERENCE   2  (bases 1 to 5922)
  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: "(2018) 
            bioRxiv 276170"
COMMENT     Sequence Label: NODAL-MYC-DYK-N72A-N199A
FEATURES             Location/Qualifiers
     source          1..5922
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     151..173
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     165..182
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     166..182
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        343..722
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        723..926
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     877..897
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     952..971
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        952..970
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2088..2117
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2136..2159
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2179..2195)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2179..2195)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    2206..2828
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2405..2424)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     rep_origin      complement(2977..3565)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3057..3076)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3894..3941
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3966..3985
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4173..4967)
                     /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             complement(4173..4967)
                     /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(4284..4303)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4894..4913
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5002..5359)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5016..5151
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5168..5359
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5316..5336
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(5361..5465)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5492..5920
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5579..5598)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5789..5810
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 aacaaaatat taacgcttac aatttccatt cgccattcag gctgcgcaac tgttgggaag
       61 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
      121 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
      181 gtgccaagct gatctataca ttgaatcaat attggcaatt agccatatta gtcattggtt
      241 atatagcata aatcaatatt ggctattggc cattgcatac gttgtatcta tatcataata
      301 tgtacattta tattggctca tgtccaatat gaccgccatg ttgacattga ttattgacta
      361 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
      421 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      481 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      541 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      601 gtccgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      661 tgaccttacg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      721 tggtgatgcg gttttggcag tacaccaatg ggcgtggata gcggtttgac tcacggggat
      781 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
      841 actttccaaa atgtcgtaat aaccccgccc cgttgacgca aatgggcggt aggcgtgtac
      901 ggtgggaggt ctatataagc agagctcgtt tagtgaaccg tcagaatttt gtaatacgac
      961 tcactatagg gcggccggga attcatcgac tggatccggt accgaggaga tctgccgccg
     1021 cgatcgccat gcacgcccac tgcctgccct tccttctgca cgcctggtgg gccctactcc
     1081 aggcgggtgc tgcgacggtg gccactgcgc tcctgcgtac gcgggggcag ccctcgtcgc
     1141 catcccctct ggcgtacatg ctgagcctct accgcgaccc gctgccgagg gcagacatca
     1201 tccgcagcct acaggcagaa gatgtggcag tggatgggca ggcctggacg tttgcttttg
     1261 acttctcctt cctgagccaa caagaggatc tggcatgggc tgagctccgg ctgcagctgt
     1321 ccagccctgt ggacctcccc actgagggct cacttgccat tgagattttc caccagccaa
     1381 agcccgacac agagcaggct tcagacagct gcttagagcg gtttcagatg gacctattca
     1441 ctgtcacttt gtcccaggtc accttttcct tgggcagcat ggttttggag gtgaccaggc
     1501 ctctctccaa gtggctgaag cgccctgggg ccctggagaa gcagatgtcc agggtagctg
     1561 gagagtgctg gccgcggccc cccacaccgc ctgccaccaa tgtgctcctt atgctctact
     1621 ccgccctctc gcaggagcag aggcagctgg gtgggtccac cttgctgtgg gaagccgaga
     1681 gctcctggcg ggcccaggag ggacagctgt cctgggagtg gggcaagagg caccgtcgac
     1741 atcacttgcc agacagaagt caactgtgtc ggaaggtcaa gttccaggtg gacttcaacc
     1801 tgatcggatg gggctcctgg atcatctacc ccaagcagta caacgcctat cgctgtgagg
     1861 gcgagtgtcc taatcctgtt ggggaggagt ttcatccgac caaccatgca tacatccaga
     1921 gtctgctgaa acgttaccag ccccaccgag tcccttccac ttgttgtgcc ccagtgaaga
     1981 ccaagccgct gagcatgctg tatgtggata atggcagagt gctcctagat caccataaag
     2041 acatgatcgt ggaagaatgt gggtgcctca cgcgtacgcg gccgctcgag cagaaactca
     2101 tctcagaaga ggatctggca gcaaatgata tcctggatta caaggatgac gacgataagg
     2161 tttaaacggc cggccgcggt catagctgtt tcctgaacag atcccgggtg gcatccctgt
     2221 gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt
     2281 gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta taatattatg
     2341 gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc
     2401 ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc
     2461 gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca
     2521 tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg
     2581 gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg
     2641 ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt aaaataacta
     2701 taccagcagg aggacgtcca gacacagcat aggctacctg gccatgccca accggtggga
     2761 catttgagtt gcttgcttgg cactgtcctc tcatgcgttg ggtccactca gtagatgcct
     2821 gttgaattgg gtacgcggcc agcggcgagc ggtatcagct cactcaaagg cggtaatacg
     2881 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     2941 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     3001 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     3061 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     3121 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     3181 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     3241 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     3301 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     3361 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     3421 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     3481 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     3541 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     3601 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     3661 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     3721 acctgaggct atggcagggc ctgccgcccc gacgttggct gcgagccctg ggccttcacc
     3781 cgaacttggg gggtggggtg gggaaaagga agaaacgcgg gcgtattggc cccaatgggg
     3841 tctcggtggg gtatcgacag agtgccagcc ctgggaccga accccgcgtt tatgaacaaa
     3901 cgacccaaca ccgtgcgttt tattctgtct ttttattgcc gtcatagcgc gggttccttc
     3961 cggtattgtc tccttccgtg tttcagttag cctcccccta gggtgggcga agaactccag
     4021 catgagatcc ccgcgctgga ggatcatcca gccggcgtcc cggaaaacga ttccgaagcc
     4081 caacctttca tagaaggcgg cggtggaatc gaaatctcgt gatggcaggt tgggcgtcgc
     4141 ttggtcggtc atttcgaacc ccagagtccc gctcagaaga actcgtcaag aaggcgatag
     4201 aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc
     4261 cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc ctgatagcga
     4321 tccgccacac ccagccggcc acagtcgatg aatccagaaa agcggccatt ttccaccatg
     4381 atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc gggcatgctc
     4441 gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc gtccagatca
     4501 tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct
     4561 tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc
     4621 atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact
     4681 tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa
     4741 ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcttgcag ttcattcagg
     4801 gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccggaac
     4861 acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc
     4921 acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg aaacgatcct
     4981 catcctgtct cttgatcgat ctttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta
     5041 cttctggaat agctcagagg ccgaggcggc ctcggcctct gcataaataa aaaaaattag
     5101 tcagccatgg ggcggagaat gggcggaact gggcggagtt aggggcggga tgggcggagt
     5161 taggggcggg actatggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc
     5221 tggggagcct ggggactttc cacacctggt tgctgactaa ttgagatgca tgctttgcat
     5281 acttctgcct gctggggagc ctggggactt tccacaccct aactgacaca cattccacag
     5341 ctggttcttt ccgcctcagg actcttcctt tttcaatatt attgaagcat ttatcagggt
     5401 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     5461 ccgcgcacat ttccccgaaa agtgccacct gacgcgccct gtagcggcgc attaagcgcg
     5521 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     5581 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     5641 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     5701 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     5761 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     5821 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     5881 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt tt
//
