| 质粒名称: | pDestTol2pA2-U6:gRNA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 63157 |
| 存储实验室: | Leonard Zon |
|
载体骨架基本信息 |
|
| 载体名称: | pDestTol2pA2 (Tol2Kit#394) |
| 载体出品公司: | -- |
| 载体抗性: | 氯霉素和氨苄青霉素 |
| 空载体大小: | 6354 bp |
| 完整质粒大小: | -- |
| 载体修饰: | 添加一个斑马鱼U6启动子，驱动一个含有BseRI位点的gRNA支架来克隆种子 |
| 载体类型: | CRISPR；网关的Tol2目标向量 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | -- |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | GAGGATCATAATCAGCCATA |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pDestTol2pA2-U6:gRNA was a gift from Leonard Zon (Addgene plasmid # 63157 ; http://n2t.net/addgene:63157 ; RRID:Addgene_63157) |
| 参考文献部分: | A CRISPR/Cas9 Vector System for Tissue-Specific Gene Disruption in Zebrafish. Ablain J, Durand EM, Yang S, Zhou Y, Zon LI. Dev Cell. 2015 Mar 4. pii: S1534-5807(15)00075-1. doi: 10.1016/j.devcel.2015.01.032. 10.1016/j.devcel.2015.01.032 PubMed 25752963 |
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 pDestTol2pA2-U6:gRNA 6352 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Destination vector for Gateway based on vector #394 from the
Tol2Kit, containing a gRNA scaffold under the control of a zebrafish
U6 promoter..
ACCESSION .
VERSION .
KEYWORDS pDestTol2pA2-U6:gRNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6352)
AUTHORS Ablain J, Durand EM, Yang S, Zhou Y, Zon LI
TITLE A CRISPR/Cas9 Vector System for Tissue-Specific Gene Disruption in
Zebrafish.
JOURNAL Dev Cell. 2015 Mar 4. pii: S1534-5807(15)00075-1. doi:
10.1016/j.devcel.2015.01.032.
PUBMED 25752963
REFERENCE 2 (bases 1 to 6352)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6352
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 20..124
/gene="bla"
/label=AmpR promoter
CDS 125..985
/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 complement(343..362)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 1156..1744
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 1645..1664
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 1898..1915
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 2032..2053
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2068..2098
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2106..2122
/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)."
primer_bind 2111..2133
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 2130..2146
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2165..2185
/label=T3
/note="T3 promoter, forward primer"
promoter 2167..2185
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind 2222..2238
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2222..2238
/label=pBluescriptSK
/note="For pBluescript vector"
polyA_signal 2910..3044
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2947..2966)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3001..3020
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
misc_RNA 3443..3518
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind 3531..3547
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 3555..3678
/gene="mutant version of attR"
/label=attR3
/bound_moiety="LR Clonase(TM)"
/note="recombination site for the Gateway(R) LR reaction"
promoter 3703..3733
/label=lac UV5 promoter
/note="E. coli lac promoter with an ""up"" mutation"
CDS 3787..4467
/codon_start=1
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
DPAY"
primer_bind complement(3853..3872)
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
CDS 4787..5092
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind 4971..4990
/label=ccdB-fwd
/note="ccdB gene, forward primer"
protein_bind complement(5133..5257)
/gene="mutant version of attR"
/label=attR4
/bound_moiety="LR Clonase(TM)"
/note="recombination site for the Gateway(R) LR reaction"
primer_bind complement(5276..5292)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 6188..6209
/label=F1ori-F
/note="F1 origin, forward primer"
ORIGIN
1 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
61 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
121 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
181 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
241 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
301 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
361 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
421 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
481 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
541 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
601 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
661 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
721 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
781 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
841 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
901 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
961 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
1021 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
1081 catgaccaaa atgccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
1141 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
1201 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
1261 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
1321 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
1381 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
1441 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
1501 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
1561 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
1621 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
1681 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
1741 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
1801 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
1861 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
1921 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
1981 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
2041 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
2101 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
2161 gcgcgcaatt aaccctcact aaagggaaca aaagctggag ctccaccgcg gtggcggccg
2221 ctctagaact agtggatctg ctgggcttgc tgaaggtagg gggtcaagaa ccagaggtgt
2281 aaagtacttg agtaatttta cttgattact gtacttaagt attatttttg gggattttta
2341 ctttacttga gtacaattaa aaatcaatac ttttactttt acttaattac atttttttag
2401 aaaaaaaagt actttttact ccttacaatt ttatttacag tcaaaaagta cttatttttt
2461 ggagatcact tcattctatt ttcccttgct attaccaaac caattgaatt gcgctgatgc
2521 ccagtttaat ttaaatgtta tttattctgc ctatgaaaat cgttttcaca ttatatgaaa
2581 ttggtcagac atgttcattg gtcctttgga agtgacgtca tgtcacatct attaccacaa
2641 tgcacagcac cttgacctgg aaattaggga aattataaca gtcaatcagt ggaagaaaat
2701 ggaggaagta tgtgattcat cagcagctgc gagcagcaca gtccaaaatc agccacagga
2761 tcaagagcac ccgtggccgt atcttcgcag atctgatcta gaggatcata atcagccata
2821 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
2881 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
2941 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
3001 gtggtttgtc caaactcatc aatgtatctt atcatgtctg gatcatcatc gatgcgtctt
3061 ttgttctggt catcaaggag gggggaatgt tctgcgcatg cctgtggggg agggagaagg
3121 acacgtcact gaaaacgtcc ctgcatcaca ccgagacacc caatcactca agccgagacc
3181 agataatttt gcatatgctt taaagtttga aaaataccac ggtaaaccct cacacaaact
3241 ctggatttga gatctttcag gttttatcag tttgcaggtt tatgtcacca tgatataggg
3301 tcagacttga tctaagggag ctgaataagt ggtttagtca ctcaccacct cccaaaaaca
3361 tacccagaag tccctggtat atatagctct ccctccagct cttggttcgg ctagcactcc
3421 tcagtgacga ggagtcagta gcgttttaga gctagaaata gcaagttaaa ataaggctag
3481 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttggtacc gtaaaacgac
3541 ggccagtgaa ttatcaacta tgtataataa agttgaacga gaaacgtaaa atgatataaa
3601 tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact gtaaaacaca
3661 acatatccag tcactatggc ggccgcatta ggcaccccag gctttacact ttatgcttcc
3721 ggctcgtata atgtgtggat tttgagttag gatccgtcga gattttcagg agctaaggaa
3781 gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca atggcatcgt
3841 aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca gaccgttcag
3901 ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt ttatccggcc
3961 tttattcaca ttcttgcccg cctgatgaat gctcatccgg aattccgtat ggcaatgaaa
4021 gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa
4081 actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca gtttctacac
4141 atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc taaagggttt
4201 attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag ttttgattta
4261 aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa atattatacg
4321 caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc
4381 ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg gcaggcgggg
4441 cgtaatctag aggatccggc ttactaaaag ccagataaca gtatgcgtat ttgcgcgctg
4501 atttttgcgg tataagaata tatactgata tgtatacccg aagtatgtca aaaagaggta
4561 tgctatgaag cagcgtatta cagtgacagt tgacagcgac agctatcagt tgctcaaggc
4621 atatatgatg tcaatatctc cggttcggta agcacaacca tgcagaatga agcccgtcgt
4681 ctgcgtgccg aacgctggaa agcggaaaat caggaaggga tggctgaggt cgcccggttt
4741 attgaaatga acggctcctt tgccgacgag aacaggggct ggtgaaatgc agtttaaggt
4801 ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga gtgatattat
4861 tgacacgccc gggcgacgga tggtgatccc cctggccagt gcacgtctgc tgtcagataa
4921 agtcccccgt gaactttacc cggtggtgca tatcggggat gaaagctggc gcatgatgac
4981 caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg atctcagcca
5041 ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat aaatgtcagg
5101 ctcccttata cacagccagt ctgcaggtcg accatagtga ctggatatgt tgtgttttac
5161 agtattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata tttatatcat
5221 tttacgtttc tcgttcaact tttctataca aagttgatag cttggcgtaa tcatggtcat
5281 agctgtttcc tgctcgaggg cccatctggc ctgtgtttca gacaccaggg agtctctgct
5341 cacgtttcct gctatttgca gcctctctat caagactaat acacctcttc ccgcatcggc
5401 tgcctgtgag aggcttttca gcactgcagg attgcttttc agccccaaaa gagctaggct
5461 tgacactaac aattttgaga atcagcttct actgaagtta aatctgaggt tttacaactt
5521 tgagtagcgt gtactggcat tagattgtct gtcttatagt ttgataatta aatacaaaca
5581 gttctaaagc aggataaaac cttgtatgca tttcatttaa tgttttttga gattaaaagc
5641 ttaaacaaga atctctagtt ttctttcttg cttttacttt tacttcctta atactcaagt
5701 acaattttaa tggagtactt ttttactttt actcaagtaa gattctagcc agatactttt
5761 acttttaatt gagtaaaatt ttccctaagt acttgtactt tcacttgagt aaaatttttg
5821 agtacttttt acacctctgt caagaactcc tggacaaacc tctgacctgt gtggaacaga
5881 gtggatatgg gtgtctgaac agatattcac gtcttttgca gatcagaggg catttctggt
5941 gttgtgcaat gacctgggtc caacaggtag gcgtactcgt atccaagagc attgttggac
6001 aggaggtagt ctccattcct ataaagagga aggaatggca ccatgtagta gccgtcattg
6061 tggccaattg gtgcattggt acgtgggtag atggaccggg gaggctgatg agttcttagc
6121 cagcgtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
6181 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
6241 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
6301 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt gg
//
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