LOCUS Exported 4268 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION NLSPUFc_VP64 in expression vector pmax. ACCESSION . VERSION . KEYWORDS pAC1358-pmax-NLSPUFc_VP64 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4268) AUTHORS Cheng AW, Jillette N, Lee P, Plaskon D, Fujiwara Y, Wang W, Taghbalout A, Wang H TITLE Casilio: a versatile CRISPR-Cas9-Pumilio hybrid for gene regulation and genomic labeling. JOURNAL Cell Res. 2016 Feb;26(2):254-7. doi: 10.1038/cr.2016.3. Epub 2016 Jan 15. PUBMED 26768771 REFERENCE 2 (bases 1 to 4268) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4268 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 138..517 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 518..721 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 672..692 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" intron 813..945 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" protein_bind 960..984 /gene="mutant version of attB" /label=attB1 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" regulatory 1002..1011 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1023..1043 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 2121..2270 /codon_start=1 /product="tetrameric repeat of the minimal activation domain of herpes simplex virus VP16 (Beerli et al., 1998)" /label=VP64 /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF DLDML" protein_bind complement(2298..2322) /gene="mutant version of attB" /label=attB2 /bound_moiety="BP Clonase(TM)" /note="recombination site for the Gateway(R) BP reaction" CDS 2375..2416 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" primer_bind complement(2442..2461) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" polyA_signal 2471..2552 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind 2496..2515 /label=SV40pA-R /note="SV40 polyA, reverse primer" rep_origin complement(2750..3337) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2830..2849) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3418..4212) /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(3529..3548) /label=Neo-F /note="Neomycin resistance gene, forward primer" primer_bind 4139..4158 /label=Neo-R /note="Neomycin resistance gene, reverse primer" ORIGIN 1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta 61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc 121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg 181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc 241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat 301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc 361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga 421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg 481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac 541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt 601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc 661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagg 721 tcgtttagtg aaccgtcaga tcactagtag ctttattgcg gtagtttatc acagttaaat 781 tgctaacgca gtcagtgctc gactgatcac aggtaagtat caaggttaca agacaggttt 841 aaggaggcca atagaaactg ggcttgtcga gacagagaag attcttgcgt ttctgatagg 901 cacctattgg tcttactgac atccactttg cctttctctc cacagggaaa aaacaattga 961 caagtttgta caaaaaagca ggctccgaat tcaccggtgc cgccaccatg gggatcctcc 1021 cccccaagaa aaagaggaag gtatctagag gccgcagccg ccttttggaa gattttcgaa 1081 acaaccggta ccccaattta caactgcggg agattgctgg acatataatg gaattttccc 1141 aagaccagca tgggtccaga ttcattcagc tgaaactgga gcgtgccaca ccagctgagc 1201 gccagcttgt cttcaatgaa atcctccagg ctgcctacca actcatggtg gatgtgtttg 1261 gtaattacgt cattcagaag ttctttgaat ttggcagtct tgaacagaag ctggctttgg 1321 cagaacggat tcgaggccac gtcctgtcat tggcactaca gatgtatggc agccgtgtta 1381 tcgagaaagc tcttgagttt attccttcag accagcagaa tgagatggtt cgggaactag 1441 atggccatgt cttgaagtgt gtgaaagatc agaatggcaa tcacgtggtt cagaaatgca 1501 ttgaatgtgt acagccccag tctttgcaat ttatcatcga cgcgtttaag ggacaggtat 1561 ttgccttatc cacacatcct tatggctgcc gagtgattca gagaatcctg gagcactgtc 1621 tccctgacca gacactccct attttagagg agcttcacca gcacacagag cagctggtac 1681 aggatcaata tggaagttat gtaatcgaac atgtactgga gcacggtcgt cctgaggata 1741 aaagcaaaat tgtagcagaa atccgaggca atgtacttgt attgagtcag cacaaatttg 1801 caaacaatgt tgtgcagaag tgtgttactc acgcctcacg tacggagcgc gctgtgctca 1861 tcgacgaggt gtgcaccatg aacgacggtc cccacagtgc cttatacacc atgatgaagg 1921 accagtatgc caactacgtg gtccagaaga tgattgacgt ggcggagcca ggccagcgga 1981 agatcgtcat gcataagatc cggccccaca tcgcaactct tcgtaagtac acctatggca 2041 agcacattct ggccaagctg gagaagtact acatgaagaa cggtgttgac ttaggggggc 2101 cggccggatc cgggcgcgcc gacgcgctgg acgatttcga tctcgacatg ctgggttctg 2161 atgccctcga tgactttgac ctggatatgt tgggaagcga cgcattggat gactttgatc 2221 tggacatgct cggctccgat gctctggacg atttcgatct cgatatgtta tatatcgatt 2281 gattaattaa gaattcgacc cagctttctt gtacaaagtg gttgatatcc agcacagtgg 2341 cggccgctcg agtctagagg gcccgcggtt cgaaggtaag cctatcccta accctctcct 2401 cggtctcgat tctacgcgta ccggttagca attgtttttt cgatgagttt ggacaaacca 2461 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat 2521 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt 2581 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg 2641 gtacttaaga gggggagacc aaagggcgag acgttaaggc ctcacgtgac atgtgagcaa 2701 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 2761 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 2821 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 2881 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 2941 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 3001 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 3061 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 3121 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 3181 acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 3241 gagttggtag ctcttgatcc ggcaaacaaa ccacgctggt agcggtggtt tttttgtttg 3301 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac 3361 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgccgtctca 3421 gaagaactcg tcaagaaggc gatagaaggc gatgcgctgc gaatcgggag cggcgatacc 3481 gtaaagcacg aggaagcggt cagcccattc gccgccaagc tcttcagcaa tatcacgggt 3541 agccaacgct atgtcctgat agcggtccgc cacacccagc cggccacagt cgatgaatcc 3601 agaaaagcgg ccattttcca ccatgatatt cggcaagcag gcatcgccat gggtcacgac 3661 gagatcctcg ccgtcgggca tgctcgcctt gagcctggcg aacagttcgg ctggcgcgag 3721 cccctgatgc tcttcgtcca gatcatcctg atcgacaaga ccggcttcca tccgagtacg 3781 tgctcgctcg atgcgatgtt tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt 3841 atgcagccgc cgcattgcat cagccatgat ggatactttc tcggcaggag caaggtgaga 3901 tgacaggaga tcctgccccg gcacttcgcc caatagcagc cagtcccttc ccgcttcagt 3961 gacaacgtcg agcacagctg cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc 4021 tgcctcgtct tgcagttcat tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg 4081 gcgcccctgc gctgacagcc ggaacacggc ggcatcagag cagccgattg tctgttgtgc 4141 ccagtcatag ccgaatagcc tctccaccca agcggccgga gaacctgcgt gcaatccatc 4201 ttgttcaatc ataatattat tgaagcattt atcagggttc gtctcgtccc ggtctcctcc 4261 catgcatg //