LOCUS Exported 2734 bp ds-DNA circular SYN 18-OCT-2021 DEFINITION Expressing paired pegRNAs from human U6 and H1 promoters to make 710-bp deletion on e-NMU locus. ACCESSION . VERSION . KEYWORDS Lentiguide-Blast-eNMU710 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 2734) TITLE Precise genomic deletions using paired prime editing JOURNAL Unpublished REFERENCE 2 (bases 1 to 2734) AUTHORS . TITLE Direct Submission JOURNAL Exported Oct 18, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..2734 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 32..272 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 32..52 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 203..222 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 302..377 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" primer_bind 429..451 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 443..460 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 444..460 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 602..621 /label=H1 /note="Human H1 promoter, forward primer" rep_origin complement(935..1523) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1015..1034) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1694..2554) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 2317..2336 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2555..2659) /gene="bla" /label=AmpR promoter ORIGIN 1 gactggatcc ggtaccaagg tcgggcagga agagggccta tttcccatga ttccttcata 61 tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac 121 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt 181 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga 241 tttcttggct ttatatatct tgtggaaagg acgaaacacc gaaggggcat gaagtttact 301 ggttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag 361 tggcaccgag tcggtgcagg tcagagtcct ggctctgtga ctcagtgata aacttcatgc 421 ctttttttcc cagtcacgac gttgtaaaac gacggccagt gaattcgaac gctgacgtca 481 tcaacccgct ccaaggaatc gcgggcccag tgtcactagg cgggaacacc cagcgcgcgt 541 gcgccctggc aggaagatgg ctgtgaggga caggggagtg gcgccctgca atatttgcat 601 gtcgctatgt gttctgggaa atcaccataa acgtgaaatg tctttggatt tgggaatctt 661 ataagttctg tatgaggacc acagatcccc agctctgtga ctcagtgacc tggaatagaa 721 aacaaaagtt taagttattc taaggccagt ccggaatcat cctaaaaagg aggcaccgag 781 tcggtgcaca tggtacccat gaaggggcat gaagtttatc actgagtcac atttttttaa 841 gcttgggccg ctcgaggtac ctctctacat atgacatgtg agcaaaaggc cagcaaaagg 901 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg 961 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat 1021 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta 1081 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct 1141 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc 1201 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa 1261 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg 1321 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag 1381 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt 1441 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta 1501 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc 1561 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca 1621 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa 1681 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat 1741 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct 1801 taccatctgg ccccagtgct gcaatgatac cgcgagatcc acgctcaccg gctccagatt 1861 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat 1921 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta 1981 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg 2041 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt 2101 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg 2161 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg 2221 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc 2281 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa 2341 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac 2401 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt 2461 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg 2521 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa 2581 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata 2641 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gctagctgta 2701 caaaaaagca ggctttaaag gaaccaattc agtc //