| 质粒名称: | pCS2TAL3-DD |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 37275 |
| 存储实验室: | David Grunwald |
|
载体骨架基本信息 |
|
| 载体名称: | pCS2 |
| 载体抗性: | 氨苄霉素 |
| 空载体大小: | 4168 bp |
| 载体修饰: | The tal1c sequence from Dan Voytas lab's pTAL3 vector (1214-2210nt) consisting of a truncated N terminal and C terminal Tal protein regions and a LacZ screening module for use in Golden Gate cloning was PCR cloned into the KpnI and BamHI sites of the pCS2-Flag-TTGZFP-FokI-DD vector from Scot Wolfe's lab, making the new vector pCS2TAL3-DD. A Esp3I site in the FokI domain of the backbone vector was modified without changing the coding sequence. |
| 载体类型: | 哺乳细胞表达，Bacterial Expression |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 启动子 | CMV |
| 标签/融合蛋白: |
Flag (N terminal on backbone)
Fok I DD (C terminal on backbone)
Tal-N (N terminal on backbone)
Tal-C (C terminal on backbone)
|
| 克隆方法: | 限制性内切酶 |
| 5' 测序引物 | TTGGCGTCGGCAAACAGTGG |
| 3' 测序引物 | GGCAACGCGATGGGACGT |
|
文献引用方法 |
|
| 材料和方法部分: | pCS2TAL3-DD was a gift from David Grunwald (爱迪基因 plasmid # 37275) |
| 参考文献部分: | Simple Methods for Generating and Detecting Locus-Specific Mutations Induced with TALENs in the Zebrafish Genome. Dahlem TJ, Hoshijima K, Jurynec MJ, Gunther D, Starker CG, Locke AS, Weis AM, Voytas DF, Grunwald DJ. PLoS Genet. 2012 Aug;8(8):e1002861. Epub 2012 Aug 16. 10.1371/journal.pgen.1002861 PubMed 22916025 |
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 pCS2TAL3-DD 5768 bp ds-DNA circular SYN 10-JAN-2018
DEFINITION TALEN backbone vector for use with the Golden Gate TALEN Kit in
place of pTAL1-4. Shorter N- and C-terminal tal protein segments
increases mutation induction; Fok I DD (improved
version=pCS2TAL3-DDD).
ACCESSION .
VERSION .
KEYWORDS pCS2TAL3-DD
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5768)
AUTHORS Dahlem TJ, Hoshijima K, Jurynec MJ, Gunther D, Starker CG, Locke AS,
Weis AM, Voytas DF, Grunwald DJ
TITLE Simple Methods for Generating and Detecting Locus-Specific Mutations
Induced with TALENs in the Zebrafish Genome.
JOURNAL PLoS Genet. 2012 Aug;8(8):e1002861. Epub 2012 Aug 16.
PUBMED 22916025
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT ORIGDB|GenBank
COMMENT VNTAUTHORNAME|biofeng.com|
COMMENT VNTNAME|pCS2TAL3-DD|
FEATURES Location/Qualifiers
source 1..5768
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 35..53
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 35..52
/label=SP6
/note="SP6 promoter, forward primer"
regulatory 85..94
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 97..117
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 118..183
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS complement(490..897)
/codon_start=1
/gene="lacZ fragment"
/product="LacZ-alpha fragment of beta-galactosidase"
/label=lacZ-alpha
/translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLQGG
TRQCIARCMHCLHCGHAATFCNLHKLACVQL"
primer_bind 747..769
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 761..778
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 762..778
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 788..807
/label=T7
/note="T7 promoter, forward primer"
promoter 788..806
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter complement(854..872)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(893..909)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 917..933
/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(941..971)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 986..1007
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
CDS 1193..1780
/codon_start=1
/product="nonspecific DNA cleavage domain of the FokI
endonuclease (Li et al., 1992)"
/label=FokI cleavage domain
/note="must dimerize to be catalytically active"
/translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV
MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQD
YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
promoter complement(1791..1808)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
polyA_signal 1813..1947
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1837..1856)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 1891..1910
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
promoter complement(2064..2082)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(2103..2119)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 2127..2143
/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(2151..2181)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2196..2217
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(2334..2351)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2505..3093)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2585..2604)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3264..4124)
/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 3887..3906
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4125..4229)
/gene="bla"
/label=AmpR promoter
rep_origin complement(4255..4710)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4392..4413)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 4604..4623
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 4784..5768
/label=CMV IE94 promoter
/note="enhancer/promoter region of simian cytomegalovirus
major immediate early transcription unit IE94"
ORIGIN
1 cgccattctg cctggggacg tcggagcaag cttgatttag gtgacactat agaatacaag
61 ctacttgttc tttttgcagg atctgccacc atggctccaa agaagaagcg taaggtagac
121 tacaaagacc atgacggtga ttataaagat catgacatcg attacaagga tgacgatgac
181 aagggtaccg tggatctacg cacgctcggc tacagtcagc agcagcaaga gaagatcaaa
241 ccgaaggtgc gttcgacagt ggcgcagcac cacgaggcac tggtgggcca tgggtttaca
301 cacgcgcaca tcgttgcgct cagccaacac ccggcagcgt tagggaccgt cgctgtcacg
361 tatcagcaca taatcacggc gttgccagag gcgacacacg aagacatcgt tggcgtcggc
421 aaacagtggt ccggcgcacg cgccctggag gccttgctca cggatgcggg ggagttgaga
481 ggtccgccgt tacagttgga cacaggccaa cttgtgaaga ttgcaaaacg tggcggcgtg
541 accgcaatgg aggcagtgca tgcatcgcgc aatgcactga cgggtgcccc cctggagacg
601 ggcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt gggaagggcg
661 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
721 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
781 gcgcgcgtaa tacgactcac tatagggcga attgggtacc gggccccccc tcgaggtcct
841 ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca tggtcatagc
901 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
961 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
1021 cactgcccgc tttccaccgg tcgtctccaa cgaccacctc gtcgccttgg cctgcctcgg
1081 cggacgtcct gccatggatg cagtgaaaaa gggattgccg cacgcgccgg aattgatcag
1141 aagagtcaat cgccgtattg gcgaacgcac gtcccatcgc gttgccggat cccaactagt
1201 caaaagtgaa ctggaggaga agaaatctga acttcgtcat aaattgaaat atgtgcctca
1261 tgaatatatt gaattaattg aaattgccag aaattccact caggatagaa ttcttgaaat
1321 gaaggtaatg gaatttttta tgaaagttta tggatataga ggtaaacatt tgggtggatc
1381 aaggaaaccg gacggagcaa tttatactgt cggatctcct attgattacg gtgtgatcgt
1441 ggatactaaa gcttatagcg gaggttataa tctgccaatt ggccaagcag atgaaatgca
1501 agactatgtc gaagaaaatc aaacacgaaa caaacatatc aaccctaatg aatggtggaa
1561 agtctatcca tcttctgtaa cggaatttaa gtttttattt gtgagtggtc actttaaagg
1621 aaactacaaa gctcagctta cacgattaaa tcatatcact aattgtaatg gagctgttct
1681 tagtgtagaa gagcttttaa ttggtggaga aatgattaaa gccggcacat taaccttaga
1741 ggaagtgcgc cggaaattta ataacggcga gataaacttt taatctagaa ctatagtgag
1801 tcgtattacg tagatccaga catgataaga tacattgatg agtttggaca aaccacaact
1861 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
1921 accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag
1981 gttcaggggg aggtgtggga ggttttttaa ttcgcggccg cggcgccaat gcattgggcc
2041 cggtacgtac ccagcttttg ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca
2101 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
2161 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
2221 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
2281 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
2341 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
2401 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
2461 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
2521 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
2581 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
2641 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
2701 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
2761 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
2821 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
2881 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
2941 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
3001 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
3061 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
3121 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
3181 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
3241 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
3301 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
3361 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
3421 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
3481 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
3541 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
3601 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
3661 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
3721 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
3781 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
3841 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
3901 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
3961 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
4021 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
4081 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
4141 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
4201 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
4261 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
4321 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
4381 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
4441 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
4501 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
4561 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
4621 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
4681 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
4741 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gtcgaccata gccaattcaa
4801 tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt gccaattcaa
4861 tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca gccaattcaa
4921 tatggcggac ttggcaccat gccaattcaa tatggcggac ttggcactgt gccaactggg
4981 gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg tgccaagtcc
5041 gccatattga attggcatgg tgccaataat ggcggccata ttggctatat gccaggatca
5101 atatataggc aatatccaat atggccctat gccaatatgg ctattggcca ggttcaatac
5161 tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt gacgtagata
5221 gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac cgcccatagc
5281 cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac gtcatatggg
5341 cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta aatggcccgc
5401 ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa tgggatggct
5461 cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg taaatggccc
5521 acttggcagt acatcaatat ctattaatag taacttggca agtacattac tattggaagg
5581 acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg gcccgcgatg
5641 gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa atgggcgttc
5701 cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc aatgctcgtt
5761 tagggaac
//
