| 质粒名称: | pAC1404-pCR8-mRuby2_NLSPUFa |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 71902 |
| 存储实验室: | Albert Cheng |
|
载体骨架基本信息 |
|
| 载体名称: | pCR8/GW/TOPO |
| 载体出品公司: | -- |
| 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 |
|
| 插入基因名称: | mRuby2_PUFa |
| 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: | mRuby（插入上的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 |
|
| 材料和方法部分: | pAC1404-pCR8-mRuby2_NLSPUFa was a gift from Albert Cheng (Addgene plasmid # 71902 ; http://n2t.net/addgene:71902 ; RRID:Addgene_71902) |
| 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
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 pAC1404-pCR8-mRuby2_NLSPUFa 4754 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION mRuby2_NLSPUFa in pCR8 gateway donor vector.
ACCESSION .
VERSION .
KEYWORDS pAC1404-pCR8-mRuby2_NLSPUFa
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4754)
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 4754)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..4754
/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 707..724
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
CDS 728..760
/codon_start=1
/product="leader peptide from bacteriophage T7 gene 10"
/label=T7 tag (gene 10 leader)
/note="promotes efficient translation in E. coli"
/translation="MASMTGGQQMG"
CDS 764..787
/codon_start=1
/product="Xpress(TM) epitope tag, including an enterokinase
recognition and cleavage site"
/label=Xpress(TM) tag
/translation="DLYDDDDK"
CDS 794..1504
/codon_start=1
/product="monomeric red fluorescent protein derived from
mRuby, with improved photophysical properties (Lam et al.,
2012)"
/label=mRuby2
/translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
AKFAGLGGGMDELYK"
CDS 1541..1561
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
protein_bind complement(2633..2732)
/gene="mutant version of attL"
/label=attL2
/bound_moiety="LR Clonase(TM)"
/note="recombination site for the Gateway(R) LR reaction"
primer_bind complement(2749..2768)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(2750..2768)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(2773..2789)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2773..2789)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
CDS 3220..4011
/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 4104..4692
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4593..4612
/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 catggtgcgg ggttctcatc atcatcatca
721 tcatggtatg gctagcatga ctggtggaca gcaaatgggt cgggatctgt acgacgatga
781 cgataaggat ccgatggtgt ctaagggcga agagctgatc aaggaaaata tgcgtatgaa
841 ggtggtcatg gaaggttcgg tcaacggcca ccaattcaaa tgcacaggtg aaggagaagg
901 caatccgtac atgggaactc aaaccatgag gatcaaagtc atcgagggag gacccctgcc
961 atttgccttt gacattcttg ccacgtcgtt catgtatggc agccgtactt ttatcaagta
1021 cccgaaaggc attcctgatt tctttaaaca gtcctttcct gagggtttta cttgggaaag
1081 agttacgaga tacgaagatg gtggagtcgt caccgtcatg caggacacca gccttgagga
1141 tggctgtctc gtttaccacg tccaagtcag aggggtaaac tttccctcca atggtcccgt
1201 gatgcagaag aagaccaagg gttgggagcc taatacagag atgatgtatc cagcagatgg
1261 tggtctgagg ggatacactc atatggcact gaaagttgat ggtggtggcc atctgtcttg
1321 ctctttcgta acaacttaca ggtcaaaaaa gaccgtcggg aacatcaaga tgcccggtat
1381 ccatgccgtt gatcaccgcc tggaaaggtt agaggaaagt gacaatgaaa tgttcgtagt
1441 acaacgcgaa cacgcagttg ccaagttcgc cgggcttggt ggtgggatgg acgagctgta
1501 caagggaggc ggcggtagcg ggccggccgg gatcctcccc cccaagaaaa agaggaaggt
1561 atctagaggc cgcagccgcc ttttggaaga ttttcgaaac aaccggtacc ccaatttaca
1621 actgcgggag attgctggac atataatgga attttcccaa gaccagcatg ggtccagatt
1681 cattcagctg aaactggagc gtgccacacc agctgagcgc cagcttgtct tcaatgaaat
1741 cctccaggct gcctaccaac tcatggtgga tgtgtttggt aattacgtca ttcagaagtt
1801 ctttgaattt ggcagtcttg aacagaagct ggctttggca gaacggattc gaggccacgt
1861 cctgtcattg gcactacaga tgtatggcag ccgtgttatc gagaaagctc ttgagtttat
1921 tccttcagac cagcagaatg agatggttcg ggaactagat ggccatgtct tgaagtgtgt
1981 gaaagatcag aatggcaatc acgtggttca gaaatgcatt gaatgtgtac agccccagtc
2041 tttgcaattt atcatcgatg cgtttaaggg acaggtattt gccttatcca cacatcctta
2101 tggctgccga gtgattcaga gaatcctgga gcactgtctc cctgaccaga cactccctat
2161 tttagaggag cttcaccagc acacagagca gctggtacag gatcaatatg gaaattatgt
2221 aatccaacat gtactggagc acggtcgtcc tgaggataaa agcaaaattg tagcagaaat
2281 ccgaggcaat gtacttgtat tgagtcagca caaatttgca agcaatgttg tggagaagtg
2341 tgttactcac gcctcacgta cggagcgcgc tgtgctcatc gatgaggtgt gcaccatgaa
2401 cgacggtccc cacagtgcct tatacaccat gatgaaggac cagtatgcca actacgtggt
2461 ccagaagatg attgacgtgg cggagccagg ccagcggaag atcgtcatgc ataagatccg
2521 gccccacatc gcaactcttc gtaagtacac ctatggcaag cacattctgg ccaagctgga
2581 gaagtactac atgaagaacg gtgttgactt agggtgatta attaagaatt cgacccagct
2641 ttcttgtaca aagttggcat tataaaaaat aattgctcat caatttgttg caacgaacag
2701 gtcactatca gtcaaaataa aatcattatt tgccatccag ctgatatccc ctatagtgag
2761 tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca aaatctctga
2821 tgttacattg cacaagataa aaatatatca tcatgcctcc tctagaccag ccaggacaga
2881 aatgcctcga cttcgctgct gcccaaggtt gccgggtgac gcacaccgtg gaaacggatg
2941 aaggcacgaa cccagtggac ataagcctgt tcggttcgta agctgtaatg caagtagcgt
3001 atgcgctcac gcaactggtc cagaaccttg accgaacgca gcggtggtaa cggcgcagtg
3061 gcggttttca tggcttgtta tgactgtttt tttggggtac agtctatgcc tcgggcatcc
3121 aagcagcaag cgcgttacgc cgtgggtcga tgtttgatgt tatggagcag caacgatgtt
3181 acgcagcagg gcagtcgccc taaaacaaag ttaaacatca tgagggaagc ggtgatcgcc
3241 gaagtatcga ctcaactatc agaggtagtt ggcgtcatcg agcgccatct cgaaccgacg
3301 ttgctggccg tacatttgta cggctccgca gtggatggcg gcctgaagcc acacagtgat
3361 attgatttgc tggttacggt gaccgtaagg cttgatgaaa caacgcggcg agctttgatc
3421 aacgaccttt tggaaacttc ggcttcccct ggagagagcg agattctccg cgctgtagaa
3481 gtcaccattg ttgtgcacga cgacatcatt ccgtggcgtt atccagctaa gcgcgaactg
3541 caatttggag aatggcagcg caatgacatt cttgcaggta tcttcgagcc agccacgatc
3601 gacattgatc tggctatctt gctgacaaaa gcaagagaac atagcgttgc cttggtaggt
3661 ccagcggcgg aggaactctt tgatccggtt cctgaacagg atctatttga ggcgctaaat
3721 gaaaccttaa cgctatggaa ctcgccgccc gactgggctg gcgatgagcg aaatgtagtg
3781 cttacgttgt cccgcatttg gtacagcgca gtaaccggca aaatcgcgcc gaaggatgtc
3841 gctgccgact gggcaatgga gcgcctgccg gcccagtatc agcccgtcat acttgaagct
3901 agacaggctt atcttggaca agaagaagat cgcttggcct cgcgcgcaga tcagttggaa
3961 gaatttgtcc actacgtgaa aggcgagatc accaaggtag tcggcaaata accctcgagc
4021 cacccatgac caaaatccct taacgtgagt tacgcgtcgt tccactgagc gtcagacccc
4081 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
4141 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
4201 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
4261 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
4321 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
4381 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
4441 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagcattga
4501 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
4561 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
4621 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
4681 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
4741 tttgctcaca tgtt
//
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