| 质粒名称: | p426*-SpSgH |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 136993 |
| 存储实验室: | Huimin Zhao |
|
载体骨架基本信息 |
|
| 载体名称: | pRS426 |
| 载体出品公司: | Zhao lab |
| 空载体大小: | 5300bp |
| 完整质粒大小: | 5294bp |
| 载体类型: | Yeast Expression |
| 筛选标记: | URA3 |
| 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 |
|
| 启动子: | SNR52 |
| 5’测序引物 | gaaagttggtgcgcatg |
| 3’测序引物 | ggaaacagctatgaccatg |
|
文献引用方法 |
|
| 材料和方法部分: | p426*-SpSgH was a gift from Huimin Zhao (Addgene plasmid # 136993 ; http://n2t.net/addgene:136993 ; RRID:Addgene_136993) |
| 参考文献部分: | Multi-functional genome-wide CRISPR system for high throughput genotype-phenotype mapping. Lian J, Schultz C, Cao M, HamediRad M, Zhao H. Nat Commun. 2019 Dec 19;10(1):5794. doi: 10.1038/s41467-019-13621-4. 10.1038/s41467-019-13621-4 PubMed 31857575 |
买家导航
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 p426*-SpSgH 5294 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Helper plasmid for SpCas9 gRNA cloning and expression.
ACCESSION .
VERSION .
KEYWORDS p426*-SpSgH
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5294)
AUTHORS Lian J, Schultz C, Cao M, HamediRad M, Zhao H
TITLE Multi-functional genome-wide CRISPR system for high throughput
genotype-phenotype mapping.
JOURNAL Nat Commun. 2019 Dec 19;10(1):5794. doi: 10.1038/s41467-019-13621-4.
PUBMED 31857575
REFERENCE 2 (bases 1 to 5294)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5294
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(44..66)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 166..185
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
promoter 209..424
/gene="S. cerevisiae URA3"
/label=URA3 promoter
CDS 425..1228
/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 1317..1585
/label=SNR52 promoter
/note="promoter for the S. cerevisiae small nucleolar RNA
gene SNR52"
misc_RNA 1602..1677
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
terminator 1689..1708
/label=SUP4 terminator
/note="transcription terminator for the S. cerevisiae SUP4
tRNA gene"
primer_bind complement(1746..1762)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1746..1762)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(1759..1781)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 1770..1786
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
promoter complement(1794..1824)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1839..1860
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(1977..1994)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2148..2736)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2228..2247)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2907..3767)
/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 3530..3549
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(3768..3872)
/gene="bla"
/label=AmpR promoter
rep_origin 3899..5241
/label=2u ori
/note="yeast 2u plasmid origin of replication"
protein_bind complement(4862..4909)
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
primer_bind 5282..6
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
ORIGIN
1 cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc
61 tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg
121 gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga
181 ttgtactgag agtgcaccat accacctttt caattcatca tttttttttt attctttttt
241 ttgatttcgg tttccttgaa atttttttga ttcggtaatc tccgaacaga aggaagaacg
301 aaggaaggag cacagactta gattggtata tatacgcata tgtagtgttg aagaaacatg
361 aaattgccca gtattcttaa cccaactgca cagaacaaaa acctgcagga aacgaagata
421 aatcatgtcg aaagctacat ataaggaacg tgctgctact catcctagtc ctgttgctgc
481 caagctattt aatatcatgc acgaaaagca aacaaacttg tgtgcttcat tggatgttcg
541 taccaccaag gaattactgg agttagttga agcattaggt cccaaaattt gtttactaaa
601 aacacatgtg gatatcttga ctgatttttc catggagggc acagttaagc cgctaaaggc
661 attatccgcc aagtacaatt ttttactctt cgaagacaga aaatttgctg acattggtaa
721 tacagtcaaa ttgcagtact ctgcgggtgt atacagaata gcagaatggg cagacattac
781 gaatgcacac ggtgtggtgg gcccaggtat tgttagcggt ttgaagcagg cggcagaaga
841 agtaacaaag gaacctagag gccttttgat gttagcagaa ttgtcatgca agggctccct
901 atctactgga gaatatacta agggtactgt tgacattgcg aagagcgaca aagattttgt
961 tatcggcttt attgctcaaa gagacatggg tggaagagat gaaggttacg attggttgat
1021 tatgacaccc ggtgtgggtt tagatgacaa gggagacgca ttgggtcaac agtatagaac
1081 cgtggatgat gtcgtttcta caggatctga cattattatt gttggaagag gactatttgc
1141 aaagggaagg gatgctaagg tagagggtga acgttacaga aaagcaggct gggaagcata
1201 tttgagaaga tgcggccagc aaaactaaaa aactgtatta taagtaaatg catgtatact
1261 aaactcacaa attagagctt caatttaatt atatcagtta ttaccctgtg ctcgagtctt
1321 tgaaaagata atgtatgatt atgctttcac tcatatttat acagaaactt gatgttttct
1381 ttcgagtata tacaaggtga ttacatgtac gtttgaagta caactctaga ttttgtagtg
1441 ccctcttggg ctagcggtaa aggtgcgcat tttttcacac cctacaatgt tctgttcaaa
1501 agattttggt caaacgctgt agaagtgaaa gttggtgcgc atgtttcggc gttcgaaact
1561 gctccgcagt gaaagataaa tgatcggaga ccgaggtctc cgttttagag ctagaaatag
1621 caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt
1681 tttgatcctt tttttgtttt ttatgtctga gctccctgca ggcatgcaag cttggcgtaa
1741 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
1801 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
1861 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
1921 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
1981 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
2041 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
2101 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
2161 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
2221 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
2281 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
2341 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
2401 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
2461 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
2521 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
2581 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
2641 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
2701 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
2761 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
2821 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
2881 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
2941 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
3001 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcggct cccacgctca
3061 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
3121 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
3181 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
3241 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
3301 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
3361 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
3421 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
3481 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
3541 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
3601 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
3661 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
3721 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
3781 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
3841 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgaa
3901 cgaagcatct gtgcttcatt ttgtagaaca aaaatgcaac gcgagagcgc taatttttca
3961 aacaaagaat ctgagctgca tttttacaga acagaaatgc aacgcgaaag cgctatttta
4021 ccaacgaaga atctgtgctt catttttgta aaacaaaaat gcaacgcgag agcgctaatt
4081 tttcaaacaa agaatctgag ctgcattttt acagaacaga aatgcaacgc gagagcgcta
4141 ttttaccaac aaagaatcta tacttctttt ttgttctaca aaaatgcatc ccgagagcgc
4201 tatttttcta acaaagcatc ttagattact ttttttctcc tttgtgcgct ctataatgca
4261 gtctcttgat aactttttgc actgtaggtc cgttaaggtt agaagaaggc tactttggtg
4321 tctattttct cttccataaa aaaagcctga ctccacttcc cgcgtttact gattactagc
4381 gaagctgcgg gtgcattttt tcaagataaa ggcatccccg attatattct ataccgatgt
4441 ggattgcgca tactttgtga acagaaagtg atagcgttga tgattcttca ttggtcagaa
4501 aattatgaac ggtttcttct attttgtctc tatatactac gtataggaaa tgtttacatt
4561 ttcgtattgt tttcgattca ctctatgaat agttcttact acaatttttt tgtctaaaga
4621 gtaatactag agataaacat aaaaaatgta gaggtcgagt ttagatgcaa gttcaaggag
4681 cgaaaggtgg atgggtaggt tatataggga tatagcacag agatatatag caaagagata
4741 cttttgagca atgtttgtgg aagcggtatt cgcaatattt tagtagctcg ttacagtccg
4801 gtgcgttttt ggttttttga aagtgcgtct tcagagcgct tttggttttc aaaagcgctc
4861 tgaagttcct atactttcta gagaatagga acttcggaat aggaacttca aagcgtttcc
4921 gaaaacgagc gcttccgaaa atgcaacgcg agctgcgcac atacagctca ctgttcacgt
4981 cgcacctata tctgcgtgtt gcctgtatat atatatacat gagaagaacg gcatagtgcg
5041 tgtttatgct taaatgcgta cttatatgcg tctatttatg taggatgaaa ggtagtctag
5101 tacctcctgt gatattatcc cattccatgc ggggtatcgt atgcttcctt cagcactacc
5161 ctttagctgt tctatatgct gccactcctc aattggatta gtctcatcct tcaatgctat
5221 catttccttt gatattggat catactaaga aaccattatt atcatgacat taacctataa
5281 aaataggcgt atca
//
