pGuide-IT-ZsGreen1

价格:8000元

联系方式:I47-825O-882O

买家导航

载体基本信息

出品公司: Clontech
载体名称: pGuide-it-ZsGreen1
质粒类型: 慢病毒载体;CRISPR/Cas9载体;荧光报告载体
高拷贝/低拷贝: 高拷贝
克隆方法: 限制性内切酶,多克隆位点
启动子: Minimal CMV
载体大小: 8303 bp
5' 测序引物及序列: --
3' 测序引物及序列: --
载体标签: --
载体抗性: 氨苄青霉素
筛选标记: 绿色荧光蛋白ZsGreen1
克隆菌株: Stbl3、 Stellar
宿主细胞(系): 常规细胞系(293、CV-1、CHO等)
备注: 慢病毒载体pGuide-it-ZsGreen1是CRISPR/Cas9载体;Clontech提供的是线性化的质粒,Biofeng提供的是环状质粒。
产品目录号: 632603
稳定性: 稳表达
组成型/诱导型: 组成型
病毒/非病毒: 慢病毒

载体质粒图谱和多克隆位点信息

pGuide-it-ZsGreen1载体图谱



pGuide-it-ZsGreen1 多克隆位点

pGuide-it-ZsGreen1 载体特征

载体简介


CRISPR/Cas9 技术

The Guide-it CRISPR/Cas9 System (Green, Cat. No. 632601; Red, Cat. No. 632602) is a complete system for the cloning and expression of a target single guide RNA (sgRNA) for mammalian gene modification studies using CRISPR/Cas9 technology. The included vector is used to simultaneously express Cas9, a target-specific sgRNA, and an exceptionally bright fluorescent protein (either ZsGreen1 or tdTomato) for monitoring transfection efficiency and/or for further enriching/isolating transfected cells by flow cytometry. The vector is pre-linearized for simple insertion of a target sequence for sgRNA expression from the human U6 promoter; Cas9 nuclease and either ZsGreen1 or tdTomato are co-expressed from a bidirectional CMV promoter. Sufficient reagents are provided in this kit for construction of 10 different target (sgRNA) expression plasmids. To construct the vector, a pair of user-provided oligos corresponding to the target genomic sequence of interest are annealed to form a duplex. Then, the duplexed DNA is cloned into the pre-linearized vector using the included high-efficiency ligation mix. This kit also contains necessary controls and Stellar? Competent Cells.

Cloning and Expression System for sgRNAs
The Guide-it CRISPR/Cas9 Systems are kits for the cloning and expression of target single guide RNAs (sgRNAs) for mammalian genome editing using CRISPR/Cas9 technology. The vector in this system simultaneously expresses Cas9 nuclease, a target-specific sgRNA, and an exceptionally bright fluorescent protein for monitoring transfection efficiency and/or for further enriching/isolating transfected cells by flow cytometry (ZsGreen1 and tdTomato versions are available). Generating a plasmid that expresses a sequence-specific sgRNA with this system is simple; a pair of user-provided oligos corresponding to the target genomic sequence of interest are annealed to form a duplex, and the duplexed oligos are inserted into the pre-linearized vector using the included high-efficiency ligation mix. The kit also includes Stellar competent cells to ensure high efficiency transformation.

Features
A single vector expresses your target-specific sgRNA from the human U6 promoter, and Cas9 nuclease and a bright fluorescent marker (either ZsGreen1 or tdTomato) are expressed from a bidirectional CMV promoter tdTomato and ZsGreen1 are exceptionally bright fluorescent proteins, 6x and 2.5x brighter than EGFP, respectively; these markers can be used to monitor transfection  efficiency and/or to enrich/isolate transfected cells by flow cytometry Simple plasmid construction—design oligos specific to your target sequence, and clone into the pre-linearized plasmid using the included ligation reagents and high- efficiency competent cells Includes sufficient reagents for construction of 10 different target (sgRNA) expression plasmids

Applications
Cloning and expression of a target single guide RNA (sgRNA) for mammalian genome editing using CRISPR/Cas9 technology

载体序列

LOCUS       pGuide-it-ZsGreen1 Vector (Linear)           8307 bp ds-DNA    linear   SYN 05-DEC-2014
COMMENT     Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT     Cat No. 632601, 632603
SOURCE      synthetic DNA construct
FEATURES             Location/Qualifiers
     misc_feature    1..93
                     /label="sgRNA"
                     /note="single-guide RNA scaffold segment; user clones guide sequence"
     misc_feature    complement(147..746)
                     /label="pUC origin"
                     /note="ori;high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(908..1768)
                     /note="AmpR"
                     /note="beta-lactamase, confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
     polyA_signal    complement(1904..2091)
                     /label="SV40 polyA signal"
     CDS             complement(2129..2821)
                     /label="ZsGreen1"
     promoter        complement(2839..2907)
                     /label="Minimal CMV Promoter 2"
     misc_feature    2971..3380
                     /label="Enhancer"
     promoter        3381..3506
                     /label="Minimal CMV Promoter 1"
     CDS             3551..7654
                     /label="Cas9"
     CDS             7670..7690
                     /label="NLS"
                     /note="nuclear localization signal of SV40 large T antigen"
     CDS             7691..7714
                     /label="DYKDDDDK tag"
     misc_feature    7753..7940
                     /label="SV40 polyA signal"
     promoter        8057..8297
                     /label="U6 Promoter"
                     /note="U6 promoter;RNA polymerase III promoter for human U6
                     snRNA"
ORIGIN
        1 gtttaagagc tatgctggaa acagcatagc aagtttaaat aaggctagtc cgttatcaac
       61 ttgaaaaagt ggcaccgagt cggtgctttt tttgagcaaa aggccagcaa aaggccagga
      121 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
      181 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
      241 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
      301 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
      361 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
      421 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
      481 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
      541 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
      601 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
      661 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
      721 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
      781 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
      841 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
      901 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
      961 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     1021 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     1081 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     1141 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     1201 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     1261 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     1321 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     1381 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     1441 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     1501 cgagttgctc ttgcccggcg tcaacacggg ataataccgc gccacatagc agaactttaa
     1561 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     1621 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     1681 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     1741 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     1801 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     1861 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcggcagt gaaaaaaatg
     1921 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     1981 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga
     2041 ggttttttaa agcaagtaaa acctctacaa atgtggtatg gctgattatg atcctctaga
     2101 ctgcagcctc aggagatctg ggccctcagg gcaaggcgga gccggaggcg atggcgtgct
     2161 cggtcaggtg ccacttctgg ttcttggcgt cgctgcggtc ctcgcgggtc agcttgtgct
     2221 ggatgaagtg ccagtcgggc atcttgcggg gcacggactt ggccttgtac acggtgtcga
     2281 actggcagcg caagcggcca ccgtccttca gcagcaggta catgctcacg tcgcccttca
     2341 agatgccctg cttgggcacg gggatgatct tctcgcagga gggctcccag ttgtcggtca
     2401 tcttcttcat cacggggccg tcggcgggga agttcacgcc gtagaacttg gactcgtggt
     2461 acatgcagtt ctcctccacg ctcacggtga tgtcggcgtt gcagatgcac acggcgccgt
     2521 cctcgaacag gaaggagcgg tcccaggtgt agccggcggg gcaggagttc ttgaagtagt
     2581 cgacgatgtc ctgggggtac tcggtgaaca cgcggttgcc gtacatgaag gcggcggaca
     2641 agatgtcctc ggcgaagggc aaggggccgc cctccaccac gcacaggttg atggcctgct
     2701 tgcccttgaa ggggtagccg atgccctcgc cggtgatcac gaacttgtgg ccgtccacgc
     2761 agccctccat gcggtacttc atggtcatct ccttggtcag gccgtgcttg gactgggcca
     2821 tggtggcgaa ttctccaggc gatctgacgg ttcactaaac gagctctgct tatataggcc
     2881 tcccaccgta cacgccacct cgacatactc gagtagttat taatagtaat caattacggg
     2941 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     3001 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     3061 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     3121 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     3181 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
     3241 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
     3301 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     3361 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
     3421 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
     3481 tggtttagtg aaccgtcaga tccgctaggg atcctctagt cagctgacgc gtgctagcgc
     3541 ggccgccacc atggacaaga agtacagcat cggcctggac atcggtacca acagcgtggg
     3601 ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aagttcaagg tgctgggcaa
     3661 caccgaccgc cacagcatca agaagaacct gatcggcgcc ctgctgttcg acagcggcga
     3721 gaccgccgag gccacccgcc tgaagcgcac cgcccgccgc cgctacaccc gccgcaagaa
     3781 ccgcatctgc tacctgcagg agatcttcag caacgagatg gccaaggtgg acgacagctt
     3841 cttccaccgc ctggaggaga gcttcctggt ggaggaggac aagaagcacg agcgccaccc
     3901 catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca
     3961 cctgcgcaag aagctggtgg acagcaccga caaggccgac ctgcgcctga tctacctggc
     4021 cctggcccac atgatcaagt tccgcggcca cttcctgatc gagggcgacc tgaaccccga
     4081 caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga
     4141 ggagaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgagcg cccgcctgag
     4201 caagagccgc cgcctggaga acctgatcgc ccagctgccc ggcgagaaga agaacggcct
     4261 gttcggcaac ctgatcgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga
     4321 cctggccgag gacgccaagc tgcagctgag caaggacacc tacgacgacg acctggacaa
     4381 cctgctggcc cagatcggcg accagtacgc cgacctgttc ctggccgcca agaacctgag
     4441 cgacgccatc ctgctgagcg acatcctgcg cgtgaacacc gagatcacca aggcccccct
     4501 gagcgccagc atgatcaagc gctacgacga gcaccaccag gacctgaccc tgctgaaggc
     4561 cctggtgcgc cagcagctgc ccgagaagta caaggagatc ttcttcgacc agagcaagaa
     4621 cggctacgcc ggctacatcg acggcggcgc cagccaggag gagttctaca agttcatcaa
     4681 gcccatcctg gagaagatgg acggcaccga ggagctgctg gtgaagctga accgcgagga
     4741 cctgctgcgc aagcagcgca ccttcgacaa cggcagcatc ccccaccaga tccacctggg
     4801 cgagctgcac gccatcctgc gccgccagga ggacttctac cccttcctga aggacaaccg
     4861 cgagaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc ccctggcccg
     4921 cggcaacagc cgcttcgcct ggatgacccg caagagcgag gagaccatca ccccctggaa
     4981 cttcgaggag gtggtggaca agggcgccag cgcccagagc ttcatcgagc gcatgaccaa
     5041 cttcgacaag aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta
     5101 cttcaccgtg tacaacgagc tgaccaaggt gaagtacgtg accgagggca tgcgcaagcc
     5161 cgccttcctg agcggcgagc agaagaaggc catcgtggac ctgctgttca agaccaaccg
     5221 caaggtgacc gtgaagcagc tgaaggagga ctacttcaag aagatcgagt gcttcgacag
     5281 cgtggagatc agcggcgtgg aggaccgctt caacgccagc ctgggcacct accacgacct
     5341 gctgaagatc atcaaggaca aggacttcct ggacaacgag gagaacgagg acatcctgga
     5401 ggacatcgtg ctgaccctga ccctgttcga ggaccgcgag atgatcgagg agcgcctgaa
     5461 gacctacgcc cacctgttcg acgacaaggt gatgaagcag ctgaagcgcc gccgctacac
     5521 cggctggggc cgcctgagcc gcaagcttat caacggcatc cgcgacaagc agagcggcaa
     5581 gaccatcctg gacttcctga agagcgacgg cttcgccaac cgcaacttca tgcagctgat
     5641 ccacgacgac agcctgacct tcaaggagga catccagaag gcccaggtga gcggccaggg
     5701 cgacagcctg cacgagcaca tcgccaacct ggccggcagc cccgccatca agaagggcat
     5761 cctgcagacc gtgaaggtgg tggacgagct ggtgaaggtg atgggccgcc acaagcccga
     5821 gaacatcgtg atcgagatgg cccgcgagaa ccagaccacc cagaagggcc agaagaacag
     5881 ccgcgagcgc atgaagcgca tcgaggaggg catcaaggag ctgggcagcc agatcctgaa
     5941 ggagcacccc gtggagaaca cccagctgca gaacgagaag ctgtacctgt actacctgca
     6001 gaacggccgc gacatgtacg tggaccagga gctggacatc aaccgcctga gcgactacga
     6061 cgtggaccac atcgtgcccc agagcttcct gaaggacgac agcatcgaca acaaggtgct
     6121 gacccgcagc gacaagaacc gcggcaagag cgacaacgtg cccagcgagg aggtggtgaa
     6181 gaagatgaag aactactggc gccagctgct gaacgccaag ctgatcaccc agcgcaagtt
     6241 cgacaacctg accaaggccg agcgcggcgg cctgagcgag ctggacaagg ccggcttcat
     6301 caagcgccag ctggtggaga cccgccagat caccaagcac gtggcccaga tcctggacag
     6361 ccgcatgaac accaagtacg acgagaacga caagctgatc cgcgaggtga aggtgatcac
     6421 cctgaagagc aagctggtga gcgacttccg caaggacttc cagttctaca aggtgcgcga
     6481 gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtggtgg gcaccgccct
     6541 gatcaagaag taccccaagc tggagagcga gttcgtgtac ggcgactaca aggtgtacga
     6601 cgtgcgcaag atgatcgcca agagcgagca ggagatcggc aaggccaccg ccaagtactt
     6661 cttctacagc aacatcatga acttcttcaa gaccgagatc accctggcca acggcgagat
     6721 ccgcaagcgc cccctgatcg agaccaacgg cgagaccggc gagatcgtgt gggacaaggg
     6781 ccgcgacttc gccaccgtgc gcaaggtgct gagcatgccc caggtgaaca tcgtgaagaa
     6841 gaccgaggtg cagaccggcg gcttcagcaa ggagagcatc ctgcccaagc gcaacagcga
     6901 caagctgatc gcccgcaaga aggactggga ccccaagaag tacggcggct tcgacagccc
     6961 caccgtggcc tacagcgtgc tggtggtggc caaggtggag aagggcaaga gcaagaagct
     7021 gaagagcgtg aaggagctgc tgggcatcac catcatggag cgcagcagct tcgagaagaa
     7081 ccccatcgac ttcctggagg ccaagggcta caaggaggtg aagaaggacc tgatcatcaa
     7141 gctgcccaag tacagcctgt tcgagctgga gaacggccgc aagcgcatgc tggccagcgc
     7201 cggcgagctg cagaagggca acgagctggc cctgcccagc aagtacgtga acttcctgta
     7261 cctggccagc cactacgaga agctgaaggg cagccccgag gacaacgagc agaagcagct
     7321 gttcgtggag cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttcag
     7381 caagcgcgtg atcctggccg acgccaacct ggacaaggtg ctgagcgcct acaacaagca
     7441 ccgcgacaag cccatccgcg agcaggccga gaacatcatc cacctgttca ccctgaccaa
     7501 cctgggcgcc cccgccgcct tcaagtactt cgacaccacc atcgaccgca agcgctacac
     7561 cagcaccaag gaggtgctgg acgccaccct gatccaccag agcatcaccg gtctgtacga
     7621 gacccgcatc gacctgagcc agctgggcgg cgacggcggc tccggacctc caaagaaaaa
     7681 gagaaaagta gactacaagg acgacgatga caagtaaatc gataagcttg tcgacgatat
     7741 ctccagagga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     7801 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     7861 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     7921 gcattttttt cactgccccg agcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     7981 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     8041 gataacgcag gaaagagagg gcctatttcc catgattcct tcatatttgc atatacgata
     8101 caaggctgtt agagagataa ttggaattaa tttgactgta aacacaaaga tattagtaca
     8161 aaatacgtga cgtagaaagt aataatttct tgggtagttt gcagttttaa aattatgttt
     8221 taaaatggac tatcatatgc ttaccgtaac ttgaaagtat ttcgatttct tggctttata
     8281 tatcttgtgg aaaggacgaa acaccgg
//

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