LOCUS Exported 5528 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pDW1775 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5528) TITLE Plant ZFN cloning vector REFERENCE 2 (bases 1 to 5528) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5528 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 36..130 /label=lambda t0 terminator /note="transcription terminator from phage lambda" primer_bind 137..156 /label=T7 /note="T7 promoter, forward primer" promoter 137..155 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" terminator 214..300 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" promoter 390..569 /label=NOS promoter /note="nopaline synthase promoter" CDS 656..676 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 677..703 /codon_start=1 /product="epitope tag from the baculovirus Autographa californica GP64 envelope fusion protein" /label=AcV5 tag /translation="SWKDASGWS" CDS 726..1313 /codon_start=1 /product="nonspecific DNA cleavage domain of the FokI endonuclease (Li et al., 1992)" /label=FokI cleavage domain /note="must dimerize to be catalytically active" /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF" terminator 1336..1588 /label=NOS terminator /note="nopaline synthase terminator and poly(A) signal" promoter complement(1630..1648) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" terminator 1650..1689 /label=fd terminator /note="central terminator from bacteriophage fd (Otsuka and Kunisawa, 1982)" CDS complement(1989..2648) /codon_start=1 /gene="S. cerevisiae HIS3" /product="imidazoleglycerol-phosphate dehydratase, required for histidine biosynthesis" /label=HIS3 /note="yeast auxotrophic marker" /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM" promoter complement(2649..2836) /gene="S. cerevisiae HIS3" /label=HIS3 promoter misc_feature 2987..3490 /label=CEN/ARS /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence" promoter 3545..3636 /gene="bla" /label=AmpR promoter rep_origin 4704..5292 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5193..5212 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5446..5463 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 gtcaacgaca tcacctggtt gcgagccttc tgaaggactc ctgttgatag atccagtaat 61 gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg cgttttttat 121 tggtgagaat caggcgtaat acgactcact atagggcgaa tggaactgga cgctcgcctg 181 tggtcgcagg gtggctcgta cgaccgcagg catcaaataa aacgaaaggc tcagtcgaaa 241 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat 301 ccgccagcgg atgagcggag aattaaggga gtcacgttat gacccccgcc gatgacgcgg 361 gacaagccgt tttacgtttg gaactgacag aaccgcaacg ttgaaggagc cactcagccg 421 ccggtttctg gagtttaatg agctaagcac atacgtcaga aaccattatt gcgcgttcaa 481 aagtcgccta aggtcactat caggtagcaa atatttcttg tcaaaaatgc tccactgacg 541 ttccataaat tcccctcggt atccaattag aaaggtggga ctcacgaaga tctaactcga 601 gaaagatatt gtatatatcg taacaatagg aggttcaaca atggcttcct cccctccaaa 661 gaaaaagaga aaggttagtt ggaaggacgc aagtggttgg tctagacccg ggaaccggtg 721 gatcccagct agtgaaatct gaattggaag agaagaaatc tgaacttaga cataaattga 781 aatatgtgcc acatgaatat attgaattga ttgaaatcgc aagaaattca actcaggata 841 gaatccttga aatgaaggtg atggagttct ttatgaaggt ttatggttat cgtggtaaac 901 atttgggtgg atcaaggaaa ccagacggag caatttatac tgtcggatct cctattgatt 961 acggtgtgat cgttgatact aaggcatatt caggaggtta taatcttcca attggtcaag 1021 cagatgaaat gcaaagatat gtcgaagaga atcaaacaag aaacaagcat atcaacccta 1081 atgaatggtg gaaagtctat ccatcttcag taacagaatt taagttcttg tttgtgagtg 1141 gtcatttcaa aggaaactac aaagctcagc ttacaagatt gaatcatatc actaattgta 1201 atggagctgt tcttagtgta gaagagcttt tgattggtgg agaaatgatt aaagctggta 1261 cattgacact tgaggaagtg agaaggaaat ttaataacgg tgagataaac ttttaatagg 1321 agctcgaatt tccccgatcg ttcaaacatt tggcaataaa gtttcttaag attgaatcct 1381 gttgccggtc ttgcgatgat tatcatataa tttctgttga attacgttaa gcatgtaata 1441 attaacatgt aatgcatgac gttatttatg agatgggttt ttatgattag agtcccgcaa 1501 ttatacattt aatacgcgat agaaaacaaa atatagcgcg caaactagga taaattatcg 1561 cgcgcggtgt catctatgtt actagatcgt gggcccacat tcctgtagaa ctcggtgaga 1621 cttttgttcc ctttagtgag ggttaattgt ccaaaaaaaa aggctccaaa aggagccttt 1681 aattgtttcg gtttatgctt aggacaccat tgattaacgc ttacaatttc ctgatgcggt 1741 attttctcct tacgcatctg tgcggtattt cacaccgcat agatccgtcg agttcaagag 1801 aaaaaaaaag aaaaagcaaa aagaaaaaag gaaagcgcgc ctcgttcaga atgacacgta 1861 tagaatgatg cattaccttg tcatcttcag tatcatactg ttcgtataaa tacttactga 1921 cattcatagc tatacatata tacacatgta tatatatcgt atgctggagc tttaaataat 1981 cggtgtcact acataagaac acctttggtg gagggaacat cgttggttcc attgggcgag 2041 gtggcttctc ttatggcaac cgcaagagcc ttgaacgcac tctcactacg gtgatggtca 2101 ttcttgcctc gcagacaatc aacgtggagg gtaattctgc ttgcctctgc aaaactttca 2161 agaaaatgcg ggatcatctc gcaagagagg tctcctactt tctccctttg caaaccaagt 2221 tcgacaactg cgtatggcct gttagaaagg tctaccaccg ctctggaaag tgcctcatcc 2281 aaaggcgcaa atcctgatcc aaaccttttt actccacgcg ccagtagagc ctctttaaaa 2341 gcctgaccga gagcaatccc gcagtcttca gtggtgtgat ggtcgtctat gtgtaagtca 2401 ccaatgcact caacgattag cgaccagccg gaatgcttgg ccagagcatg tatcatgtgg 2461 tccagaaacc ctatacctgt gtggacgtta atcacttgcg attgtgtggc ctgttctgct 2521 actgcttctg cctctttttc tgggaagatc gagtgctcta tcgctagggg accacccttt 2581 aaagagatag caatctgaat cttggtttca tttgtaatac gctttactag ggctttctgc 2641 tctgtcatct ttgccttcgt ttatcttgcc tgctcatttt ttagtatatt cttcgaagaa 2701 atcacattac tttatataat gtataattca ttatgtgata atgccaatcg ctaagaaaaa 2761 aaaagagtca tccgctaggt ggaaaaaaaa aaatgaaaat cattaccgag gcataaaaaa 2821 atatagagtg tactagagga ggccaagagt aatagaaaaa gaaaattgcg ggaaaggact 2881 gtgttatgac ttccctgact aatgccgtgt tcaaacgata cctggcagtg actcctagcg 2941 ctcaccaagc tcttaaaacg ggaatttatg gatcttaatt aagacggatc gcttgcctgt 3001 aacttacacg cgcctcgtat cttttaatga tggaataatt tgggaattta ctctgtgttt 3061 atttattttt atgttttgta tttggatttt agaaagtaaa taaagaaggt agaagagtta 3121 cggaatgaag aaaaaaaaat aaacaaaggt ttaaaaaatt tcaacaaaaa gcgtacttta 3181 catatatatt tattagacaa gaaaagcaga ttaaatagat atacattcga ttaacgataa 3241 gtaaaatgta aaatcacagg attttcgtgt gtggtcttct acacagacaa gatgaaacaa 3301 ttcggcatta atacctgaga gcaggaagag caagataaaa ggtagtattt gttggcgatc 3361 cccctagagt cttttacatc ttcggaaaac aaaaactatt ttttctttaa tttctttttt 3421 tactttctat ttttaattta tatatttata ttaaaaaatt taaattataa ttatttttat 3481 agcacgtgat gaaaagcctt agggtcaggt ggcacttttc ggggaaatgt gcgatgaacc 3541 cctatttgtt tatttttcta aatacattca aatatgtatc cgctaatgag acaataaccc 3601 tgataaatgc ttcaattata ttgaaaaagg aagagtatga gtatccaaca tttccgtgtc 3661 gcccttattc ccttttttgc ggccttctgc cttcctgttt ttgctcaccc agaaacgctg 3721 gtgaaagtaa aagatgctga agatcagttg ggtgcccgag tgggttacat tgaactggat 3781 ctcaacagcg gcaaaatctt agagagtttt cgccccgaag aacgcttccc aatgatgagc 3841 actttcaaag ttctgctatg tggcgcggtg ttatcccgta ttgatgctgg gcaagagcaa 3901 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtattcacc agtcacagaa 3961 aagcatctta cggatggcat gacagtaaga gaattatgta gtgctgccat aaccatgagt 4021 gataacactg cggccaactt acttctgaca acgattggag gaccgaagga gctaaccgct 4081 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat 4141 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaactctc 4201 cgcaaactat taactggcga actacttact cttgcttcac gccagcaact cattgactgg 4261 atggaggcgg acaaagttgc aggaccactt ctgcgctcgg cacttccggc tggctggttt 4321 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg 4381 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg 4441 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg 4501 tcagacgaag ggaaataaat agtagcccgc ctgatgtgcg ggcttttttt tggtggtggt 4561 taccagtgtc gcttagctac cgtaggtcct cttaggcggt ccgattagcg acatagtcat 4621 caatgccttg acggcaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc 4681 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg 4741 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctactaact 4801 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg 4861 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg 4921 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac 4981 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgctca 5041 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga 5101 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc 5161 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct 5221 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg 5281 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct 5341 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc 5401 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc 5461 gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat 5521 tgatgcag //