LOCUS Exported 5428 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Yeast Centromere Plasmid for galactose inducible expression of retron-guide chimeric RNA with flanking HH-HDV ribozyme . ACCESSION . VERSION . KEYWORDS pZS165 CRISPEY HH-HDV NotI SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5428) AUTHORS Sharon E, Chen SA, Khosla NM, Smith JD, Pritchard JK, Fraser HB TITLE Functional Genetic Variants Revealed by Massively Parallel Precise Genome Editing. JOURNAL Cell. 2018 Sep 18. pii: S0092-8674(18)31118-8. doi: 10.1016/j.cell.2018.08.057. PUBMED 30245013 REFERENCE 2 (bases 1 to 5428) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5428 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 127..231 /gene="bla" /label=AmpR promoter CDS 232..1092 /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 complement(450..469) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 1263..1851 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 1752..1771 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind complement(2950..2969) /label=T7 /note="T7 promoter, forward primer" promoter complement(2951..2969) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2979..2996) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(2979..2995) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2988..3010) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 3140..3595 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3227..3246) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3437..3458 /label=F1ori-F /note="F1 origin, forward primer" CDS complement(3726..4529) /codon_start=1 /gene="S. cerevisiae URA3" /product="orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis" /label=URA3 /note="yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)" /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN" promoter complement(4530..4745) /gene="S. cerevisiae URA3" /label=URA3 promoter primer_bind complement(4776..4795) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 4895..4917 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(4955..4973) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" misc_feature 5015..90 /label=CEN/ARS /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence" ORIGIN 1 ttttctttaa tttctttttt tactttctat ttttaattta tatatttata ttaaaaaatt 61 taaattataa ttatttttat agcacgtgat gaaaaggacc caggtggcac ttttcgggga 121 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc 181 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt 241 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct 301 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt 361 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt 421 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac 481 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac 541 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct 601 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg 661 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg 721 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca 781 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa 841 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt 901 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc 961 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg 1021 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt 1081 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt 1141 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc 1201 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct 1261 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta 1321 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc 1381 ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac 1441 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct 1501 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat 1561 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg 1621 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa 1681 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg 1741 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga 1801 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc 1861 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcaagt agagggggta 1921 atttttcccc tttattttgt tcatacattc ttaaattgct ttgcctctcc ttttggaaag 1981 ctatacttcg gagcactgtt gagcgaaggc tcattagata tattttctgt cattttcctt 2041 aacccaaaaa taagggaaag ggtccaaaaa gcgctcggac aactgttgac cgtgatccga 2101 aggactggct atacagtgtt cacaaaatag ccaagctgaa aataatgtgt agctatgttc 2161 agttagtttg gctagcaaag atataaaagc aggtcggaaa tatttatggg cattattatg 2221 cagagcatca acatgataaa aaaaaacagt tgaatattcc ctcaaaaggg tgcgcatctg 2281 atgagtccgt gaggacgaaa cgagctagct cgtcatgcgc acccttagcg agaggtttat 2341 cattaaggtc aacctctgga tgttgtttcg gcatcctgca ttgaatctga gttactgtct 2401 gttttcctgc ggccgcgttt cagagctatg ctggaaacag catagcaagt tgaaataagg 2461 ctagtccgtt atcaacttga aaaagtggca ccgagtcggt gctttttgat ggccggcatg 2521 gtcccagcct cctcgctggc gccggctggg caacaccttc gggtggcgaa tgggacttaa 2581 agaaagtgga atattcattc atatcatatt ttttctatta actgcctggt ttcttttaaa 2641 ttttttattg gttgtcgact tgaacggagt gacaatatat atatatatat atttaataat 2701 gacatcatta tctgtaaatc tgattcttaa tgctattcta gttatgtaag agtggtcctt 2761 tccataaaaa aaaaaaaaaa gaaaaaagaa ttttaggaat acaatgcagc ttgtaagtaa 2821 aatctggaat attcatatcg ccacaacttc ttatgcttat aaaagcacta atgcctgaat 2881 ttatgttgaa aatatgtgtc acaaataaag aaactgtgac atctgacaca tttccacgta 2941 cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt 3001 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc 3061 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc 3121 ctgaatggcg aatggcgcga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg 3181 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc 3241 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc 3301 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt 3361 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag 3421 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg 3481 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag 3541 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aatttcctga 3601 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcataggg taataactga 3661 tataattaaa ttgaagctct aatttgtgag tttagtatac atgcatttac ttataataca 3721 gttttttagt tttgctggcc gcatcttctc aaatatgctt cccagcctgc ttttctgtaa 3781 cgttcaccct ctaccttagc atcccttccc tttgcaaata gtcctcttcc aacaataata 3841 atgtcagatc ctgtagagac cacatcatcc acggttctat actgttgacc caatgcgtct 3901 cccttgtcat ctaaacccac accgggtgtc ataatcaacc aatcgtaacc ttcatctctt 3961 ccacccatgt ctctttgagc aataaagccg ataacaaaat ctttgtcgct cttcgcaatg 4021 tcaacagtac ccttagtata ttctccagta gatagggagc ccttgcatga caattctgct 4081 aacatcaaaa ggcctctagg ttcctttgtt acttcttctg ccgcctgctt caaaccgcta 4141 acaatacctg ggcccaccac accgtgtgca ttcgtaatgt ctgcccattc tgctattctg 4201 tatacacccg cagagtactg caatttgact gtattaccaa tgtcagcaaa ttttctgtct 4261 tcgaagagta aaaaattgta cttggcggat aatgccttta gcggcttaac tgtgccctcc 4321 atggaaaaat cagtcaagat atccacatgt gtttttagta aacaaatttt gggacctaat 4381 gcttcaacta actccagtaa ttccttggtg gtacgaacat ccaatgaagc acacaagttt 4441 gtttgctttt cgtgcatgat attaaatagc ttggcagcaa caggactagg atgagtagca 4501 gcacgttcct tatatgtagc tttcgacatg atttatcttc gtttcctgca ggtttttgtt 4561 ctgtgcagtt gggttaagaa tactgggcaa tttcatgttt cttcaacact acatatgcgt 4621 atatatacca atctaagtct gtgctccttc cttcgttctt ccttctgttc ggagattacc 4681 gaatcaaaaa aatttcaaag aaaccgaaat caaaaaaaag aataaaaaaa aaatgatgaa 4741 ttgaattgaa aagctgtggt atggtgcact ctcagtacaa tctgctctga tgccgcatag 4801 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc 4861 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt 4921 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag 4981 gttaatgtca tgataataat ggtttcttag gacggatcgc ttgcctgtaa cttacacgcg 5041 cctcgtatct tttaatgatg gaataatttg ggaatttact ctgtgtttat ttatttttat 5101 gttttgtatt tggattttag aaagtaaata aagaaggtag aagagttacg gaatgaagaa 5161 aaaaaaataa acaaaggttt aaaaaatttc aacaaaaagc gtactttaca tatatattta 5221 ttagacaaga aaagcagatt aaatagatat acattcgatt aacgataagt aaaatgtaaa 5281 atcacaggat tttcgtgtgt ggtcttctac acagacaaga tgaaacaatt cggcattaat 5341 acctgagagc aggaagagca agataaaagg tagtatttgt tggcgatccc cctagagtct 5401 tttacatctt cggaaaacaa aaactatt //