| 质粒名称: | pICE-FLAG-NAT10-siR-WT |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 59365 |
| 存储实验室: | Steve Jackson |
|
载体骨架基本信息 |
|
| 载体名称: | pICE |
| 空载体大小: | 5044bp |
| 完整质粒大小: | 8086 bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | Puromycin |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | XL1 Blue |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | human NAT10 / NAT10 / ALP /KIAA1709 |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 951 bp |
| 基因突变: | Mutations to render cDNA resistant to a siRNA |
| GenBank编号 | NM_024662 |
| 启动子: | CMV-tet |
| 标签/融合蛋白: | FLAG (N terminal on backbone) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | BamHI(未破坏) |
| 3’克隆位点: | MluI(未破坏) |
| 5’测序引物 | CMVpromoter forward |
| 3’测序引物 | BGHpolyA reverse |
|
文献引用方法 |
|
| 材料和方法部分: | pICE-FLAG-NAT10-siR-WT was a gift from Steve Jackson (Addgene plasmid # 59365 ; http://n2t.net/addgene:59365 ; RRID:Addgene_59365) |
| 参考文献部分: | Chemical inhibition of NAT10 corrects defects of laminopathic cells. Larrieu D, Britton S, Demir M, Rodriguez R, Jackson SP. Science. 2014 May 2;344(6183):527-32. doi: 10.1126/science.1252651. 10.1126/science.1252651 PubMed 24786082 |
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 pICE-FLAG-NAT10-siR-WT 8086 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Plasmid for constitutive or doxycycline-inducible expression of
wild-type human NAT10 resistant to a siRNA. Confers resistance to
puromycin. Use T-REx cells for doxycycline-inducible expression..
ACCESSION .
VERSION .
KEYWORDS pICE-FLAG-NAT10-siR-WT
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8086)
AUTHORS Larrieu D, Britton S, Demir M, Rodriguez R, Jackson SP
TITLE Chemical inhibition of NAT10 corrects defects of laminopathic cells.
JOURNAL Science. 2014 May 2;344(6183):527-32. doi: 10.1126/science.1252651.
PUBMED 24786082
REFERENCE 2 (bases 1 to 8086)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..8086
/organism="synthetic DNA construct"
/mol_type="other DNA"
regulatory 16..25
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 25..48
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
regulatory 694..703
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
primer_bind complement(3167..3184)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 3173..3397
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 3443..3871
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3530..3549)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 3740..3761
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(3880..3900)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 3885..4214
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 4065..4200
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 4127..4146
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 4266..4865
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind complement(4266..4285)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 4762..4782
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
polyA_signal 4995..5116
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(5032..5051)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 5086..5105
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(5166..5182)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5166..5182)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5179..5201)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5190..5206
/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(5214..5244)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5259..5280
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5397..5414)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5568..6153)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5648..5667)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6324..7184)
/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 6947..6966
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7185..7289)
/gene="bla"
/label=AmpR promoter
enhancer 7361..7740
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 7741..7944
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 7895..7915
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
protein_bind 7946..7964
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
protein_bind 7967..7985
/gene="tetO"
/label=tet operator
/bound_moiety="tetracycline repressor TetR"
/note="bacterial operator O2 for the tetR and tetA genes"
primer_bind 7995..8019
/label=LNCX
/note="Human CMV promoter, forward primer"
ORIGIN
1 aagcttgcgg ccgccgccac catggattac aaggatgacg acgataaggg atccatgcat
61 cggaaaaagg tggataaccg aatccggatt ctcattgaga atggagtagc tgagcggcaa
121 agatctctct ttgttgtagt tggggatcga ggaaaagatc aggtggtaat acttcatcac
181 atgttatcca aagcaactgt gaaggctcgg ccttcagtgc tgtggtgtta taagaaagag
241 ctggggttta gcagtcaccg gaagaaaaga atgcgacagc tgcagaagaa aataaagaat
301 ggaacactga acataaagca ggacgacccc tttgaactct tcatagcagc cacaaacatt
361 cgctactgct actacaacga gacccacaag atcctgggca ataccttcgg catgtgtgtg
421 ctgcaggatt ttgaagcctt aactccaaac ttgctggcca ggactgtaga aacagtggaa
481 ggtggtgggc tagtggtcat cctcctacgg accatgaact cactcaagca attgtacaca
541 gtgactatgg atgtgcattc caggtacaga actgaggccc atcaggatgt ggtgggaaga
601 tttaatgaaa ggtttattct gtctctggcc tcttgtaaga agtgtctcgt cattgatgac
661 cagctcaaca tcctgcccat ctcctcccac gttgccacca tggaggccct gcctccccag
721 actccggatg agagtcttgg tccttctgat ctggagctga gggagttgaa ggagagcttg
781 caggacaccc agcctgtggg tgtgttggtg gactgctgta agactctaga ccaggccaaa
841 gctgtcttga aatttatcga gggcatctct gaaaagaccc tgaggagtac tgttgcactc
901 acagctgctc gaggacgggg aaaatctgca gccctgggat tggcgattgc tggggcggtg
961 gcatttgggt actccaatat ctttgttacc tccccaagcc ctgataacct ccatactctg
1021 tttgaatttg tatttaaagg atttgatgct ctgcaatatc aggaacatct ggattatgag
1081 attatccagt ctctaaatcc tgaatttaac aaagcagtga tcagagtgaa tgtatttcga
1141 gaacacaggc agactattca gtatatacat cctgcagatg ctgtgaagct gggccaggct
1201 gaactagttg tgattgatga agctgccgcc atccccctcc ccttggtgaa gagcctactt
1261 ggcccctacc ttgttttcat ggcatccacc atcaatggct atgagggcac tggccggtca
1321 ctgtccctca agctaattca gcagctccgt caacagagcg cccagagcca ggtcagcacc
1381 actgctgaga ataagaccac gacgacagcc agattggcat cagcgcggac actgcatgag
1441 gtttccctcc aggagtcaat ccgatacgcc cctggggatg cagtggagaa gtggctgaat
1501 gacttgctgt gcctggattg cctcaacatc actcggatag tctcaggctg ccccttgcct
1561 gaagcttgtg aactgtacta tgttaataga gataccctct tttgctacca caaggcctct
1621 gaagttttcc tccaacggct tatggccctc tacgtggctt ctcactacaa gaactctccc
1681 aatgatctcc agatgctctc cgatgcacct gctcaccatc tcttctgcct tctgcctcct
1741 gtgcccccca cccagaatgc ccttccagaa gtgcttgctg ttatccaggt gtgccttgaa
1801 ggggagattt ctcgccagtc catcttgaac agtctgtctc gaggcaagaa ggcttcaggg
1861 gacctgattc catggacagt gtcagaacag ttccaagatc cagactttgg tggtctgtct
1921 ggtggaaggg tcgttcgcat tgctgttcac ccagattatc aagggatggg ctatggcagc
1981 cgtgctctgc agctgctgca gatgtactat gaaggcaggt ttccttgtct ggaggaaaag
2041 gtccttgaga caccacagga aattcacacc gtaagcagcg aggctgtcag cttgttggaa
2101 gaggtcatca ctccccggaa ggacctgcct cctttactcc tcaaattgaa tgagaggcct
2161 gccgaacgcc tggattacct gggtgtttcc tatggcttga cccccaggct cctcaagttc
2221 tggaaacgag ctggatttgt tcctgtttat ctgagacaga ccccgaatga cctgaccgga
2281 gagcactcgt gcatcatgct gaagacgctc actgatgagg atgaggctga ccagggaggc
2341 tggcttgcag ccttctggaa agatttccga cggcggttcc tagccttgct ctcctaccag
2401 ttcagtacct tctctccttc cctggctctg aacatcattc agaacaggaa catggggaag
2461 ccagcccagc ctgccctgag ccgggaggag ctggaagcac tcttcctccc ctatgacctg
2521 aagcggctgg agatgtattc acggaatatg gtggactatc acctcatcat ggacatgatc
2581 ccggccatct ctcgcatcta tttcctgaac cagctggggg acctggccct gtctgcggct
2641 cagtcggctc ttctcttggg gattggcctg cagcataagt ctgtggacca gctggaaaag
2701 gagattgagc tgccctcggg ccagttgatg ggacttttca accggatcat ccgcaaagtt
2761 gtgaagctat ttaatgaagt tcaggaaaag gccattgagg agcagatggt ggcagcgaag
2821 gatgtggtca tggagcccac gatgaagacc ctcagtgacg acctagatga agcagcaaag
2881 gaatttcagg agaaacacaa gaaggaagta gggaagctga aatcaatgga tttgagtgaa
2941 tatattatcc gtggggacga tgaagagtgg aatgaagttt tgaacaaagc tgggccgaac
3001 gcctcgatca tcagcctgaa aagtgacaag aaaaggaagt tagaggccaa acaagaaccc
3061 aaacagagca agaagttgaa gaacagagag acaaagaaca aaaaagatat gaaactgaag
3121 cggaagaaat agacgcgtct gcagtctaga taaacccgct gatcagcctc gactgtgcct
3181 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
3241 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
3301 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
3361 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
3421 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
3481 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
3541 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
3601 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
3661 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
3721 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
3781 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
3841 gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt
3901 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
3961 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
4021 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
4081 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
4141 ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
4201 taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg
4261 tgaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg
4321 ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc
4381 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc
4441 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc
4501 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga
4561 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca
4621 aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc
4681 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct
4741 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca
4801 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg
4861 cctgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
4921 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
4981 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
5041 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
5101 tcatcaatgt atcttatcat gtctggctag ccgtcgacct ctagctagag cttggcgtaa
5161 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
5221 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
5281 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
5341 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
5401 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
5461 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
5521 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
5581 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
5641 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
5701 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
5761 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
5821 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
5881 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
5941 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
6001 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
6061 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtttttt tgtttgcaag
6121 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
6181 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
6241 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
6301 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
6361 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
6421 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
6481 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
6541 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
6601 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
6661 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
6721 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
6781 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
6841 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
6901 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
6961 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
7021 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
7081 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
7141 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
7201 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
7261 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
7321 gacggatcgg gagatcctgt acgggccaga tatccgcgtt gacattgatt attgactagt
7381 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
7441 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
7501 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
7561 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
7621 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
7681 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
7741 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
7801 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
7861 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
7921 tgggaggtct atataagcag agctctccct atcagtgata gagatctccc tatcagtgat
7981 agagatcgtc gacgagctcg tttagtgaac cgtcagatcg cctggagacg ccatccacgc
8041 tgttttgacc tccatagaag acaccgggac cgatccagcc tccggc
//
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