质粒名称: | pAC1403-pCR8-Clover_NLSPUFc |
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出品公司: | Addgene |
目录编号: | 71901 |
存储实验室: | Albert Cheng |
载体骨架基本信息 |
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载体名称: | pCR8/GW/TOPO |
载体出品公司: | Invitrogen |
空载体大小: | -- |
完整质粒大小: | -- |
载体修饰: | -- |
载体类型: | Gateway Donor |
筛选标记: | -- |
载体抗性: | 大观霉素50 μg/mL |
生长温度: | 37 ℃ |
宿主菌: | DH5a |
拷贝数: | -- |
插入基因信息 |
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插入基因名称: | Clover_PUFc |
物种来源: | Synthetic |
插入大小: | -- |
基因突变: | -- |
GenBank编号 | -- |
启动子: | -- |
标签/融合蛋白: | Clover(插件上的N端子) |
克隆方法: | -- |
5’克隆位点: | -- |
3’克隆位点: | -- |
5’测序引物 | M13F |
3’测序引物 | M13R |
文献引用方法 |
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材料和方法部分: | pAC1403-pCR8-Clover_NLSPUFc was a gift from Albert Cheng (Addgene plasmid # 71901 ; http://n2t.net/addgene:71901 ; RRID:Addgene_71901) |
参考文献部分: | Casilio: a versatile CRISPR-Cas9-Pumilio hybrid for gene regulation and genomic labeling. Cheng AW, Jillette N, Lee P, Plaskon D, Fujiwara Y, Wang W, Taghbalout A, Wang H. Cell Res. 2016 Feb;26(2):254-7. doi: 10.1038/cr.2016.3. Epub 2016 Jan 15. 10.1038/cr.2016.3 PubMed 26768771 |
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载体基本信息
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载体简介
载体序列
LOCUS pAC1403-pCR8-Clover_NLSPUFc 4694 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Clover_NLSPUFc in pCR8 gateway donor vector. ACCESSION . VERSION . KEYWORDS pAC1403-pCR8-Clover_NLSPUFc SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4694) AUTHORS Cheng AW, Jillette N, Lee P, Plaskon D, Fujiwara Y, Wang W, Taghbalout A, Wang H TITLE Casilio: a versatile CRISPR-Cas9-Pumilio hybrid for gene regulation and genomic labeling. JOURNAL Cell Res. 2016 Feb;26(2):254-7. doi: 10.1038/cr.2016.3. Epub 2016 Jan 15. PUBMED 26768771 REFERENCE 2 (bases 1 to 4694) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4694 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 92..109 /label=L4440 /note="L4440 vector, forward primer" terminator 268..295 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" terminator 387..473 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 522..544 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 536..553 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 537..553 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" protein_bind 569..668 /gene="mutant version of attL" /label=attL1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" regulatory 686..695 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 692..1408 /codon_start=1 /product="bright green-yellow fluorescent protein derived from GFP (Lam et al., 2012)" /label=Clover /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" primer_bind complement(737..758) /label=EGFP-N /note="EGFP, reverse primer" primer_bind 1345..1366 /label=EGFP-C /note="EGFP, forward primer" promoter complement(1424..1442) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(1425..1442) /label=SP6 /note="SP6 promoter, forward primer" CDS 1481..1501 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" protein_bind complement(2573..2672) /gene="mutant version of attL" /label=attL2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" primer_bind complement(2689..2708) /label=T7 /note="T7 promoter, forward primer" promoter complement(2690..2708) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2713..2729) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2713..2729) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" CDS 3160..3951 /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK" rep_origin 4044..4632 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 4533..4552 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga 61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga 121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca 181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc 241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta 301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc 361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa 421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg 481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa 541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac 601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtacaaaaa 661 agcaggctcc gaattcaccg gtgccgccac catggtgagc aagggcgagg agctgttcac 721 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt 781 ccgcggcgag ggcgagggcg atgccaccaa cggcaagctg accctgaagt tcatctgcac 841 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcgtggc 901 ctgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc 961 cgaaggctac gtccaggagc gcaccatctc tttcaaggac gacggtacct acaagacccg 1021 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga 1081 cttcaaggag gacggcaaca tcctggggca caagctggag tacaacttca acagccacaa 1141 cgtctatatc acggccgaca agcagaagaa cggcatcaag gctaacttca agatccgcca 1201 caacgttgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg 1261 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc catcagtccg ccctgagcaa 1321 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat 1381 tacacatggc atggacgagc tgtacaagtc tagagggccc tattctatag tgtcacctaa 1441 atgcggaggc ggcggtagcg ggccggccgg gatcctcccc cccaagaaaa agaggaaggt 1501 atctagaggc cgcagccgcc ttttggaaga ttttcgaaac aaccggtacc ccaatttaca 1561 actgcgggag attgctggac atataatgga attttcccaa gaccagcatg ggtccagatt 1621 cattcagctg aaactggagc gtgccacacc agctgagcgc cagcttgtct tcaatgaaat 1681 cctccaggct gcctaccaac tcatggtgga tgtgtttggt aattacgtca ttcagaagtt 1741 ctttgaattt ggcagtcttg aacagaagct ggctttggca gaacggattc gaggccacgt 1801 cctgtcattg gcactacaga tgtatggcag ccgtgttatc gagaaagctc ttgagtttat 1861 tccttcagac cagcagaatg agatggttcg ggaactagat ggccatgtct tgaagtgtgt 1921 gaaagatcag aatggcaatc acgtggttca gaaatgcatt gaatgtgtac agccccagtc 1981 tttgcaattt atcatcgacg cgtttaaggg acaggtattt gccttatcca cacatcctta 2041 tggctgccga gtgattcaga gaatcctgga gcactgtctc cctgaccaga cactccctat 2101 tttagaggag cttcaccagc acacagagca gctggtacag gatcaatatg gaagttatgt 2161 aatcgaacat gtactggagc acggtcgtcc tgaggataaa agcaaaattg tagcagaaat 2221 ccgaggcaat gtacttgtat tgagtcagca caaatttgca aacaatgttg tgcagaagtg 2281 tgttactcac gcctcacgta cggagcgcgc tgtgctcatc gacgaggtgt gcaccatgaa 2341 cgacggtccc cacagtgcct tatacaccat gatgaaggac cagtatgcca actacgtggt 2401 ccagaagatg attgacgtgg cggagccagg ccagcggaag atcgtcatgc ataagatccg 2461 gccccacatc gcaactcttc gtaagtacac ctatggcaag cacattctgg ccaagctgga 2521 gaagtactac atgaagaacg gtgttgactt agggtgatta attaagaatt cgacccagct 2581 ttcttgtaca aagttggcat tataaaaaat aattgctcat caatttgttg caacgaacag 2641 gtcactatca gtcaaaataa aatcattatt tgccatccag ctgatatccc ctatagtgag 2701 tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca aaatctctga 2761 tgttacattg cacaagataa aaatatatca tcatgcctcc tctagaccag ccaggacaga 2821 aatgcctcga cttcgctgct gcccaaggtt gccgggtgac gcacaccgtg gaaacggatg 2881 aaggcacgaa cccagtggac ataagcctgt tcggttcgta agctgtaatg caagtagcgt 2941 atgcgctcac gcaactggtc cagaaccttg accgaacgca gcggtggtaa cggcgcagtg 3001 gcggttttca tggcttgtta tgactgtttt tttggggtac agtctatgcc tcgggcatcc 3061 aagcagcaag cgcgttacgc cgtgggtcga tgtttgatgt tatggagcag caacgatgtt 3121 acgcagcagg gcagtcgccc taaaacaaag ttaaacatca tgagggaagc ggtgatcgcc 3181 gaagtatcga ctcaactatc agaggtagtt ggcgtcatcg agcgccatct cgaaccgacg 3241 ttgctggccg tacatttgta cggctccgca gtggatggcg gcctgaagcc acacagtgat 3301 attgatttgc tggttacggt gaccgtaagg cttgatgaaa caacgcggcg agctttgatc 3361 aacgaccttt tggaaacttc ggcttcccct ggagagagcg agattctccg cgctgtagaa 3421 gtcaccattg ttgtgcacga cgacatcatt ccgtggcgtt atccagctaa gcgcgaactg 3481 caatttggag aatggcagcg caatgacatt cttgcaggta tcttcgagcc agccacgatc 3541 gacattgatc tggctatctt gctgacaaaa gcaagagaac atagcgttgc cttggtaggt 3601 ccagcggcgg aggaactctt tgatccggtt cctgaacagg atctatttga ggcgctaaat 3661 gaaaccttaa cgctatggaa ctcgccgccc gactgggctg gcgatgagcg aaatgtagtg 3721 cttacgttgt cccgcatttg gtacagcgca gtaaccggca aaatcgcgcc gaaggatgtc 3781 gctgccgact gggcaatgga gcgcctgccg gcccagtatc agcccgtcat acttgaagct 3841 agacaggctt atcttggaca agaagaagat cgcttggcct cgcgcgcaga tcagttggaa 3901 gaatttgtcc actacgtgaa aggcgagatc accaaggtag tcggcaaata accctcgagc 3961 cacccatgac caaaatccct taacgtgagt tacgcgtcgt tccactgagc gtcagacccc 4021 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg 4081 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact 4141 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg 4201 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg 4261 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac 4321 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca 4381 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagcattga 4441 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc 4501 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct 4561 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg 4621 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct 4681 tttgctcaca tgtt //