质粒名称: | pROS16 |
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出品公司: | Addgene |
目录编号: | 107930 |
存储实验室: | Jean-Marc Daran |
载体骨架基本信息 |
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载体名称: | p426-SNR52p-gRNA.CAN1.Y-SUP4t |
载体出品公司: | George Church (Addgene plasmid # 43803) |
空载体大小: | 5462 bp |
完整质粒大小: | 5465 bp |
载体类型: | Yeast Expression, CRISPR |
筛选标记: | HIS3 |
载体抗性: | 氨苄霉素 |
生长温度: | 37 ℃ |
宿主菌: | DH5a |
拷贝数: | 高拷贝 |
插入基因信息 |
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插入基因名称: |
gRNA-CAN1.Y, gRNA-ADE2.Y
gRNA/shRNA sequenceGATACGTTCTCTATGGAGGA, ACTTGAAGATTCTTTAGTGT
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物种来源: | S. cerevisiae (budding yeast) |
插入大小: | 20 bp |
克隆方法: | Gibson Cloning |
文献引用方法 |
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材料和方法部分: |
pROS16 was a gift from Jean-Marc Daran (Addgene plasmid # 107930 ; http://n2t.net/addgene:107930 ; RRID:Addgene_107930)
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参考文献部分: |
CRISPR/Cas9: a molecular Swiss army knife for simultaneous introduction of multiple genetic modifications in Saccharomyces cerevisiae. Mans R, van Rossum HM, Wijsman M, Backx A, Kuijpers NG, van den Broek M, Daran-Lapujade P, Pronk JT, van Maris AJ, Daran JM. FEMS Yeast Res. 2015 Mar;15(2). pii: fov004. doi: 10.1093/femsyr/fov004. Epub 2015 Mar 4. 10.1093/femsyr/fov004 PubMed 25743786
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载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pROS16 5465 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION HIS3 based Yeast/E.coli shuttle vector designed to clone and express Cas9 programming spacer. ACCESSION . VERSION . KEYWORDS pROS16 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5465) AUTHORS Mans R, van Rossum HM, Wijsman M, Backx A, Kuijpers NG, van den Broek M, Daran-Lapujade P, Pronk JT, van Maris AJ, Daran JM TITLE CRISPR/Cas9: a molecular Swiss army knife for simultaneous introduction of multiple genetic modifications in Saccharomyces cerevisiae. JOURNAL FEMS Yeast Res. 2015 Mar;15(2). pii: fov004. doi: 10.1093/femsyr/fov004. Epub 2015 Mar 4. PUBMED 25743786 REFERENCE 2 (bases 1 to 5465) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.biofeng.com FEATURES Location/Qualifiers source 1..5465 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(351..368) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(522..1110) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(602..621) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(1281..2141) /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 1904..1923 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2142..2246) /gene="bla" /label=AmpR promoter primer_bind 2317..2336 /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" promoter 2483..2669 /gene="S. cerevisiae HIS3" /label=HIS3 promoter CDS 2670..3332 /codon_start=1 /gene="S. cerevisiae HIS3" /product="imidazoleglycerol-phosphate dehydratase, required for histidine biosynthesis" /label=HIS3 /note="yeast auxotrophic marker" /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM" promoter 3657..3925 /label=SNR52 promoter /note="promoter for the S. cerevisiae small nucleolar RNA gene SNR52" misc_RNA 3946..4020 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" terminator 4025..4044 /label=SUP4 terminator /note="transcription terminator for the S. cerevisiae SUP4 tRNA gene" rep_origin 4269..5250 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 5296..5315 /label=SUP4 terminator /note="transcription terminator for the S. cerevisiae SUP4 tRNA gene" misc_RNA complement(5320..5394) /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter complement(5415..218) /label=SNR52 promoter /note="promoter for the S. cerevisiae small nucleolar RNA gene SNR52" ORIGIN 1 caactttcac ttctacagcg tttgaccaaa atcttttgaa cagaacattg tagggtgtga 61 aaaaatgcgc acctttaccg ctagcccaag agggcactac aaaatctaga gttgtacttc 121 aaacgtacat gtaatcacct tgtatatact cgaaagaaaa catcaagttt ctgtataaat 181 atgagtgaaa gcataatcat acattatctt ttcaaagaag ctccagcttt tgttcccttt 241 agtgagggta ttcacgtaga cggataggta tagccagaca tcagcagcat acttcgggaa 301 ccgtaggctc tagagagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact 361 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta 421 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag 481 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 541 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 601 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 661 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc 721 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac 781 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac 841 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg 901 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga 961 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt 1021 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag 1081 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct 1141 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg 1201 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat 1261 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc 1321 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg 1381 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct 1441 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca 1501 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg 1561 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg 1621 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc 1681 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag 1741 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg 1801 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag 1861 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat 1921 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg 1981 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca 2041 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca 2101 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat 2161 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag 2221 aaaaataaac aaataggggt tccgcgcact cgagactata tgtgaaggca tggctatggc 2281 acggcagaca ttccgccaga tcatcaatag gcactgcggc atcagagcag attgtactga 2341 gagtgcacca taaattcccg ttttaagagc ttggtgagcg ctaggagtca ctgccaggta 2401 tcgtttgaac acggcattag tcagggaagt cataacacag tcctttcccg caattttctt 2461 tttctattac tcttggcctc ctctagtaca ctctatattt ttttatgcct cggtaatgat 2521 tttcattttt tttttccacc tagcggatga ctcttttttt ttcttagcga ttggcattat 2581 cacataatga attatacatt atataaagta atgtgatttc ttcgaagaat atactaaaaa 2641 atgagcaggc aagataaacg aaggcaaaga tgacagagca gaaagcccta gtaaagcgta 2701 ttacaaatga aaccaagatt cagattgcga tctctttaaa gggtggtccc ctagcgatag 2761 agcactcgat cttcccagaa aaagaggcag aagcagtagc agaacaggcc acacaatcgc 2821 aagtgattaa cgtccacaca ggtatagggt ttctggacca tatgatacat gctctggcca 2881 agcattccgg ctggtcgcta atcgttgagt gcattggtga cttacacata gacgaccatc 2941 acaccactga agactgcggg attgctctcg gtcaagcttt taaagaggcc ctaggggccg 3001 tgcgtggagt aaaaaggttt ggatcaggat ttgcgccttt ggatgaggca ctttccagag 3061 cggtggtaga tctttcgaac aggccgtacg cagttgtcga acttggtttg caaagggaga 3121 aagtaggaga tctctcttgc gagatgatcc cgcattttct tgaaagcttt gcagaggcta 3181 gcagaattac cctccacgtt gattgtctgc gaggcaagaa tgatcatcac cgtagtgaga 3241 gtgcgttcaa ggctcttgcg gttgccataa gagaagccac ctcgcccaat ggtaccaacg 3301 atgttccctc caccaaaggt gttcttatgt agtgacaccg attatttaaa gctgcagcat 3361 acgatatata tacatgtgta tatatgtata cctatgaatg tcagtaagta tgtatacgaa 3421 cagtatgata ctgaagatga caaggtaatg catcattcta tacgtgtcat tctgaacgag 3481 gcgcgctttc cttttttctt tttgcttttt cttttttttt ctcttgaact cgacggatct 3541 atgcggtgtg aaataccgca cagatgcacc tttcgagagg acgatgcccg tgtctaaatg 3601 attcgaccag cctaagaatg ttcaacccct cactaaaggg aacaaaagct ggagcttctt 3661 tgaaaagata atgtatgatt atgctttcac tcatatttat acagaaactt gatgttttct 3721 ttcgagtata tacaaggtga ttacatgtac gtttgaagta caactctaga ttttgtagtg 3781 ccctcttggg ctagcggtaa aggtgcgcat tttttcacac cctacaatgt tctgttcaaa 3841 agattttggt caaacgctgt agaagtgaaa gttggtgcgc atgtttcggc gttcgaaact 3901 tctccgcagt gaaagataaa tgatcacttg aagattcttt agtgtgtttt agagctagaa 3961 atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg 4021 gtgctttttt tgttttttat gtcttcgagt catgtaatta gttatgtcac gcttacgttc 4081 acgccctcca cgcatttaag cataaacacg cactatgccg ttcttctcat gtatatatat 4141 atacaggcaa cacgcagata taggtgcgac gtgaacagtg agctgtatgt gcgcagctcg 4201 cgttgcattt tcggaagcgc tcgttttcgg aaacgctttg aagttcctat tccgaagttc 4261 ctattctgta gaaagtatag gaacctcaga gcgcttttga aaaccaaaag cgctctgaag 4321 acgcactttc aaaaaaccaa aaacgcaccg gactgtaacg agctactaaa atattgcgaa 4381 taccgcttcc acaaacattg ctcaaaagta tctctttgct atatatctct gtgctatatc 4441 cctatataac ctacccatcc acctttcgct ccttgaactt gcatctaaac tcgacctcta 4501 cattttttat gtttatctct agtattactc tttagacaaa aaaattgtag taagaactat 4561 tcatagagtg aatcgaaaac aatacgaaaa tgtaaacatt tcctatacgt agtatataga 4621 gacaaaatag aagaaaccgt tcataatttt ctgaccaatg aagaatcatc aacgctatca 4681 ctttctgttc acaaagtatg cgcaatccac atcggtatag aatataatcg gggatgcctt 4741 tatcttgaaa aaatgcaccc gcagcttcgc tagtaatcag taaacgcggg aagtggagtc 4801 aggctttttt tatggaagag aaaatagaca ccaaagtagc cttcttctaa ccttaacgga 4861 cctacagtgc aaaaagttat caagagactg cattatagag cgcacaaagg agaaaaaaag 4921 taatctaaga tgctttgtta gaaaaatagc gctctcggga tgcatttttg tagaacaaaa 4981 aagaagtata gattctttgt tggtaaaata gcgctctcgc gttgcatttc tgttctgtaa 5041 aaatgcagct cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat ttttgtttta 5101 caaaaatgaa gcacagattc ttcgttggta aaatagcgct ttcgcgttgc atttctgttc 5161 tgtaaaaatg cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg 5221 ttctacaaaa tgaagcacag atgcttcgtt ggagggcgtg aacgtaagcg tgacataact 5281 aattacatga ctcgaagaca taaaaaacaa aaaaagcacc accgactcgg tgccactttt 5341 tcaagttgat aacggactag ccttatttta acttgctatt tctagctcta aaactcctcc 5401 atagagaacg tatcgatcat ttatctttca ctgcggagaa gtttcgaacg ccgaaacatg 5461 cgcac //
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