| 质粒名称: | pcDNA3/Flag-WTAP |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 53741 |
| 存储实验室: | Chuan He |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3 |
| 载体出品公司: | Invitrogen |
| 空载体大小: | 5446bp |
| 完整质粒大小: | 6530 bp |
| 载体类型: | 哺乳细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 未知 |
|
插入基因信息 |
|
| 插入基因名称: | WTAP |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 1191bp |
| GenBank编号 | NM_001270531.1 |
| 启动子: | T7 |
| 标签/融合蛋白: | FLAG tag (N terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | AgeI(未破坏) |
| 3’克隆位点: | XhoI (未破坏) |
| 5’测序引物 | 5' TAATACGACTCACTATAGGG 3' |
| 3’测序引物 | 5' ATTTAGGTGACACTATAG 3' |
|
文献引用方法 |
|
| 材料和方法部分: |
pcDNA3/Flag-WTAP was a gift from Chuan He (Addgene plasmid # 53741 ; http://n2t.net/addgene:53741 ; RRID:Addgene_53741)
|
| 参考文献部分: | A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine methylation. Liu J, Yue Y, Han D, Wang X, Fu Y, Zhang L, Jia G, Yu M, Lu Z, Deng X, Dai Q, Chen W, He C. Nat Chem Biol. 2014 Feb;10(2):93-5. doi: 10.1038/nchembio.1432. Epub 2013 Dec 6. 10.1038/nchembio.1432 PubMed 24316715 |
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 pcDNA3/Flag-WTAP 6530 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION For expression of WTAP (Wilms tumor 1 associated protein) in human
cells.
ACCESSION .
VERSION .
KEYWORDS pcDNA3/Flag-WTAP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6530)
AUTHORS Liu J, Yue Y, Han D, Wang X, Fu Y, Zhang L, Jia G, Yu M, Lu Z, Deng
X, Dai Q, Chen W, He C
TITLE A METTL3-METTL14 complex mediates mammalian nuclear RNA N6-adenosine
methylation.
JOURNAL Nat Chem Biol. 2014 Feb;10(2):93-5. doi: 10.1038/nchembio.1432. Epub
2013 Dec 6.
PUBMED 24316715
REFERENCE 2 (bases 1 to 6530)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..6530
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 107..128
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 143..173
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 181..197
/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 186..208
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 205..221
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 205..221
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
polyA_signal 270..391
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(281..300)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 335..354
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
CDS complement(565..1359)
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(676..695)
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind 1286..1305
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
promoter complement(1426..1622)
/label=SV40 promoter
/note="SV40 early promoter"
promoter complement(1434..1644)
/label=SV40 promoter
/note="SV40 early promoter"
rep_origin 1440..1575
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind complement(1494..1513)
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind 1668..1688
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin complement(1697..2125)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1807..1828)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 2019..2038
/label=F1ori-R
/note="F1 origin, reverse primer"
polyA_signal 2171..2395
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind 2384..2401
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
promoter 2421..2439
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 2421..2438
/label=SP6
/note="SP6 promoter, forward primer"
CDS complement(3655..3678)
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
primer_bind complement(3712..3731)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(3713..3731)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter complement(3776..3979)
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind complement(3805..3825)
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
enhancer 3980..4359
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
primer_bind 4531..4550
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
promoter 4625..4729
/gene="bla"
/label=AmpR promoter
CDS 4730..5590
/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(4948..4967)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5761..6349
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6250..6269
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 6503..6520
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct
241 agctagaggt cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac
301 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
361 tgtaaccatt ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat
421 ccccgcgctg gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt
481 catagaaggc ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg
541 tcatttcgaa ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat
601 gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc
661 gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac
721 acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg
781 caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatgc gcgccttgag
841 cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc
901 gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc
961 gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga
1021 tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa
1081 tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc
1141 cgtcgtggcc agccacgata gccgcgctgc ctcgtcctgc agttcattca gggcaccgga
1201 caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc
1261 atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc
1321 ggccggagaa cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc ctcatcctgt
1381 ctcttgatca gatccgaaaa tggatataca agctcccggg agctttttgc aaaagcctag
1441 gcctccaaaa aagcctcctc actacttctg gaatagctca gaggcagagg cggcctcggc
1501 ctctgcataa ataaaaaaaa ttagtcagcc atggggcgga gaatgggcgg aactgggcgg
1561 agttaggggc gggatgggcg gagttagggg cgggactatg gttgctgact aattgagatg
1621 catgctttgc atacttctgc ctgctgggga gcctggggac tttccacacc ctaactgaca
1681 cacattccac agaattaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
1741 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
1801 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
1861 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
1921 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
1981 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
2041 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
2101 cttaatgcgc cgctacaggg cgcgtgggga taccccctag agccccagct ggttctttcc
2161 gcctcagaag ccatagagcc caccgcatcc ccagcatgcc tgctattgtc ttcccaatcc
2221 tcccccttgc tgtcctgccc caccccaccc cccagaatag aatgacacct actcagacaa
2281 tgcgatgcaa tttcctcatt ttattaggaa aggacagtgg gagtggcacc ttccagggtc
2341 aaggaaggca cgggggaggg gcaaacaaca gatggctggc aactagaagg cacagtcgag
2401 gctgatcagc gagctctagc atttaggtga cactatagaa tagggccctc tagatgcatg
2461 ctcgagttac aaaactgaac cctgtacatt tacacttgag tccaagccat tctgaacgtg
2521 gcgggaaccc acagttcgat tacctttccc actcactgct ttctcctctt gagggtcatg
2581 actggagtct gtgtcatttg attggtgtgt gagagagttt tcactgcccg tgggagagtc
2641 tacactttca taccccgcac tgagttgatt tacgtaacca ctacctcctc tgccagttct
2701 ctcctctgag ctgttggagg acttattacc attccctgga gaagaaggaa agtcatcttc
2761 ggttgtgttg ccctccctgt gaaatccaga cccagacgtc ctctggcggg aggagctacc
2821 attacttggt ccgtttgtca gacgactgca gtctttactt gtggcctctg actgttctac
2881 aggttcagaa gctgtagtcc tgctggtact tggggcagag gcctgagact gctgctgctg
2941 gtactgagcc aactgctggc gtgtctcctt cagctgctgc tgcagaacta gaatggtact
3001 ctgcataccc tctacttctt catcaagctg gatgatgaag tcattcagtt catcctgact
3061 gcttttaagc tcctcactgt atttcttctg taaagccaac tctgcttcaa gttgtgcaat
3121 acgtccctgg gacagctgcc ttccaagctc ttgattctcc tggataagca ttcgacactt
3181 cgccattaac tttttccctg tttggctatc aggcgtaaac ttccaggcac tcagttcatt
3241 ttgggcttgt tccagtttgt ctttagtctg ttccagttca cctttcattt ttaggaaaaa
3301 caagttgatc gctgggtcta ccattgttga tctcagttgg gcaacgctcg gctgctggac
3361 ttgcttgagg tactggattt gagtagtaca ctcttgcatc tcttgttcct tggttgctag
3421 tcgcattaca aggatgtttt ccctgcgtgc agactcctgc tgttgttgct ttagtttttc
3481 ttcagactct cttaggccag ttacatcatt agagttaaga tctgtgtact tgccctccaa
3541 agcttgtaca tatgcttcat attgtttcca tcttagaatt aactcatctc ttgccataac
3601 tttgaagtct gtttcactca atcgaacctt cttgggaaga ggttcttcgt tggtcttgtc
3661 atcgtcatcc ttgtaatcca tggatccgag ctcggtacca agcttgggtc tccctatagt
3721 gagtcgtatt aatttcgata agccagtaag cagtgggttc tctagttagc cagagagctc
3781 tgcttatata gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt
3841 tgttacgaca ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt
3901 caatggggtg gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt
3961 actgccaaaa ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag
4021 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
4081 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
4141 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
4201 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
4261 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
4321 taattgatta ctattaataa ctagtcaata atcaatgtca acgcgtatat ctggcccgta
4381 catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt cttcatgcaa ttgtcggtca
4441 agccttgcct tgttgtagct taaattttgc tcgcgcacta ctcagcgacc tccaacacac
4501 aagcagggag cagatactgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
4561 accatagggg atcgggagat ctcccgatcc gtcgacgtca ggtggcactt ttcggggaaa
4621 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
4681 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
4741 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
4801 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
4861 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
4921 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
4981 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
5041 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
5101 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
5161 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
5221 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
5281 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
5341 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
5401 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
5461 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
5521 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
5581 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
5641 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
5701 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
5761 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
5821 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
5881 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
5941 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
6001 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
6061 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
6121 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
6181 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
6241 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
6301 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
6361 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
6421 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
6481 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
//
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