LOCUS Exported 3266 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION pMAGIC L3-L2 entry plasmid, contains 3xHA tag+polyA; empty hU6-driven xCas9(3.7) gRNA scaffold for 3-/4-component MultiSite Gateway Pro assembly. Allows C-term HA fusion to dCas9 w/ gRNA expression. ACCESSION . VERSION . KEYWORDS KK701: pMAGIC (L3-L2) 3x HA eptitope tag + polyA; hU6::xCas9(3... SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3266) AUTHORS Haldeman JM, Conway AE, Arlotto ME, Slentz DH, Muoio DM, Becker TC, Newgard CB TITLE Creation of versatile cloning platforms for transgene expression and dCas9-based epigenome editing. JOURNAL Nucleic Acids Res. 2018 Dec 27. pii: 5264291. doi: 10.1093/nar/gky1286. PUBMED 30590691 REFERENCE 2 (bases 1 to 3266) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3266 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 92..109 /label=L4440 /note="L4440 vector, forward primer" terminator 268..295 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" terminator 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=attL3 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS 677..757 /codon_start=1 /product="three tandem HA epitope tags" /label=3xHA /translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA" primer_bind complement(785..802) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 791..998 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" promoter 1005..1245 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 1005..1025 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 1176..1195 /label=LKO.1 5' /note="Human U6 promoter, forward primer" protein_bind complement(1378..1477) /gene="mutant version of attL" /label=attL2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" primer_bind complement(1507..1526) /label=T7 /note="T7 promoter, forward primer" promoter complement(1508..1526) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1531..1547) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1531..1547) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1649..1668) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 1660..2469 /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(1727..1746) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin 2616..3204 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3105..3124 /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 tgtataataa 661 agttggtggc ggatcctacc catacgatgt tccagattac gcttacccat acgatgttcc 721 agattacgct tacccatacg atgttccaga ttacgcttaa gaattcctag agctcgctga 781 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct 841 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca 901 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag 961 ggggaggatt gggaagagaa tagcaggcat gctggggagg taccgagggc ctatttccca 1021 tgattccttc atatttgcat atacgataca aggctgttag agagataatt ggaattaatt 1081 tgactgtaaa cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg 1141 ggtagtttgc agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt 1201 gaaagtattt cgatttcttg gctttatata tcttgtggaa aggacgaaac accggagacc 1261 acggcaggtc tcagtttcag agctatgctg gaaacagcat agcaagttga aataaggcta 1321 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct tttttgcggc cgcaggaacc 1381 cagctttctt gtacaaagtt ggcattataa gaaagcattg cttatcaatt tgttgcaacg 1441 aacaggtcac tatcagtcaa aataaaatca ttatttgcca tccagctgca gggcggccgc 1501 gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca gctctggccc 1561 gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc atgaacaata 1621 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa 1681 acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta taaatgggct 1741 cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa gcccgatgcg 1801 ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac agatgagatg 1861 gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca ttttatccgt 1921 actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc attccaggta 1981 ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt gttcctgcgc 2041 cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt atttcgtctc 2101 gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt tgatgacgag 2161 cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt gccattctca 2221 ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt tgacgagggg 2281 aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata ccaggatctt 2341 gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg gctttttcaa 2401 aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat gctcgatgag 2461 tttttctaat cagaattggt taattggttg taacactggc agagcattac gctgacttga 2521 cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc gttccactga 2581 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta 2641 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa 2701 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact 2761 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca 2821 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt 2881 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg 2941 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag 3001 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta 3061 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat 3121 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg 3181 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc 3241 ttttgctggc cttttgctca catgtt //