LOCUS Exported 5955 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION For mammalian expression of the human NODAL open reading frame (with a mutated Cysteine residue) with an internal MYC tag. ACCESSION . VERSION . KEYWORDS NODAL-matMYC-C312S SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5955) 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 bioRxiv 276170 REFERENCE 2 (bases 1 to 5955) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5955 /organism="synthetic DNA construct" /mol_type="other DNA" 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 1743..1772 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 2121..2150 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 2169..2192 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" primer_bind complement(2212..2228) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2212..2228) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" polyA_signal 2239..2861 /label=hGH poly(A) signal /note="human growth hormone polyadenylation signal" primer_bind complement(2438..2457) /label=hGH-PA-R /note="Human growth hormone terminator, reverse primer" rep_origin complement(3010..3598) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3090..3109) /label=pBR322ori-F /note="pBR322 origin, forward primer" polyA_signal 3927..3974 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" primer_bind 3999..4018 /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" CDS complement(4206..5000) /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(4317..4336) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 4927..4946 /label=Neo-R /note="Neomycin resistance gene, reverse primer" promoter complement(5035..5392) /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 5049..5184 /label=SV40 ori /note="SV40 origin of replication" primer_bind complement(5103..5122) /label=SV40pro-F /note="SV40 promoter/origin, forward primer" primer_bind 5349..5369 /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter complement(5394..5498) /gene="bla" /label=AmpR promoter rep_origin 5525..25 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(5612..5631) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 5822..5843 /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 gaactggacg 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 caccctgggg ccctggagaa gcagatgtcc agggtagctg 1561 gagagtgctg gccgcggccc cccacaccgc ctgccaccaa tgtgctcctt atgctctact 1621 ccaacctctc gcaggagcag aggcagctgg gtgggtccac cttgctgtgg gaagccgaga 1681 gctcctggcg ggcccaggag ggacagctgt cctgggagtg gggcaagagg caccgtcgac 1741 atgagcagaa actcatctca gaagaggatc tgcatcactt gccagacaga agtcaactgt 1801 gtcggaaggt caagttccag gtggacttca acctgatcgg atggggctcc tggatcatct 1861 accccaagca gtacaacgcc tatcgctgtg agggcgagtg tcctaatcct gttggggagg 1921 agtttcatcc gaccaaccat gcatacatcc agagtctgct gaaacgttac cagccccacc 1981 gagtcccttc cacttcctgt gccccagtga agaccaagcc gctgagcatg ctgtatgtgg 2041 ataatggcag agtgctccta gatcaccata aagacatgat cgtggaagaa tgtgggtgcc 2101 tctgacgtac gcggccgctc gagcagaaac tcatctcaga agaggatctg gcagcaaatg 2161 atatcctgga ttacaaggat gacgacgata aggtttaaac ggccggccgc ggtcatagct 2221 gtttcctgaa cagatcccgg gtggcatccc tgtgacccct ccccagtgcc tctcctggcc 2281 ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag ttgcatcatt 2341 ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt atggagcaag 2401 gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc aagctggagt 2461 gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc gattctcctg 2521 cctcagcctc ccgagttgtt gggattccag gcatgcatga ccaggctcag ctaatttttg 2581 tttttttggt agagacgggg tttcaccata ttggccaggc tggtctccaa ctcctaatct 2641 caggtgatct acccaccttg gcctcccaaa ttgctgggat tacaggcgtg aaccactgct 2701 cccttccctg tccttctgat tttaaaataa ctataccagc aggaggacgt ccagacacag 2761 cataggctac ctggccatgc ccaaccggtg ggacatttga gttgcttgct tggcactgtc 2821 ctctcatgcg ttgggtccac tcagtagatg cctgttgaat tgggtacgcg gccagcggcg 2881 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 2941 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 3001 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 3061 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 3121 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 3181 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 3241 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 3301 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 3361 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 3421 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 3481 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 3541 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 3601 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 3661 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 3721 aagttttaaa tcaatctaaa gtatatatga gtaacctgag gctatggcag ggcctgccgc 3781 cccgacgttg gctgcgagcc ctgggccttc acccgaactt ggggggtggg gtggggaaaa 3841 ggaagaaacg cgggcgtatt ggccccaatg gggtctcggt ggggtatcga cagagtgcca 3901 gccctgggac cgaaccccgc gtttatgaac aaacgaccca acaccgtgcg ttttattctg 3961 tctttttatt gccgtcatag cgcgggttcc ttccggtatt gtctccttcc gtgtttcagt 4021 tagcctcccc ctagggtggg cgaagaactc cagcatgaga tccccgcgct ggaggatcat 4081 ccagccggcg tcccggaaaa cgattccgaa gcccaacctt tcatagaagg cggcggtgga 4141 atcgaaatct cgtgatggca ggttgggcgt cgcttggtcg gtcatttcga accccagagt 4201 cccgctcaga agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg 4261 gcgataccgt aaagcacgag gaagcggtca gcccattcgc cgccaagctc ttcagcaata 4321 tcacgggtag ccaacgctat gtcctgatag cgatccgcca cacccagccg gccacagtcg 4381 atgaatccag aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccatgg 4441 gtcacgacga gatcctcgcc gtcgggcatg ctcgccttga gcctggcgaa cagttcggct 4501 ggcgcgagcc cctgatgctc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc 4561 cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga 4621 tcaagcgtat gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca 4681 aggtgagatg acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc 4741 gcttcagtga caacgtcgag cacagctgcg caaggaacgc ccgtcgtggc cagccacgat 4801 agccgcgctg cctcgtcttg cagttcattc agggcaccgg acaggtcggt cttgacaaaa 4861 agaaccgggc gcccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc 4921 tgttgtgccc agtcatagcc gaatagcctc tccacccaag cggccggaga acctgcgtgc 4981 aatccatctt gttcaatcat gcgaaacgat cctcatcctg tctcttgatc gatctttgca 5041 aaagcctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc 5101 ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga 5161 actgggcgga gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta 5221 attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct 5281 ggttgctgac taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga 5341 ctttccacac cctaactgac acacattcca cagctggttc tttccgcctc aggactcttc 5401 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt 5461 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca 5521 cctgacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg 5581 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc 5641 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga 5701 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt 5761 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat 5821 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat 5881 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa 5941 tttaacgcga atttt //