| 质粒名称: | METTL3 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 155725 |
| 存储实验室: | Eugene Yeo |
|
载体骨架基本信息 |
|
| 载体名称: | E2.pEF5-DEST-FL-V5-MCP |
| 空载体大小: | 7357 bp |
| 完整质粒大小: | 8110 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: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | METTL3 / Accession: BC052244 |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 1743bp |
| 基因突变: | H330Q compared to BC052244, no mutations compared to ABZ92354.1 |
| GenBank编号 | ABZ92354.1 |
| 启动子: | EF-1a |
| 标签/融合蛋白: | V5-MCP (C terminal on backbone) |
| 克隆方法: | Gateway Cloning |
| 5’测序引物 | TAATACGACTCACTATAGGG |
| 3’测序引物 | gctcgagtctagagggc |
|
文献引用方法 |
|
| 材料和方法部分: | TTL3 was a gift from Eugene Yeo (Addgene plasmid # 155725 ; http://n2t.net/addgene:155725 ; RRID:Addgene_155725) |
| 参考文献部分: | Large-scale tethered function assays identify factors that regulate mRNA stability and translation. Luo EC, Nathanson JL, Tan FE, Schwartz JL, Schmok JC, Shankar A, Markmiller S, Yee BA, Sathe S, Pratt GA, Scaletta DB, Ha Y, Hill DE, Aigner S, Yeo GW. Nat Struct Mol Biol. 2020 Oct;27(10):989-1000. doi: 10.1038/s41594-020-0477-6. Epub 2020 Aug 17. 10.1038/s41594-020-0477-6 PubMed 32807991 |
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 METTL3 8110 bp ds-DNA circular SYN 08-NOV-2022
DEFINITION For use in RBP tethering screen.
ACCESSION .
VERSION .
KEYWORDS METTL3
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8110)
TITLE Yeo Lab plasmids
REFERENCE 2 (bases 1 to 8110)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Nov 8, 2022 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..8110
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..1179
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
intron 232..1170
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind 1127..1147
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
primer_bind 1196..1215
/label=T7
/note="T7 promoter, forward primer"
promoter 1196..1214
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 1293..1317
/gene="mutant version of attB"
/label=attB1
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
protein_bind complement(3067..3091)
/gene="mutant version of attB"
/label=attB2
/bound_moiety="BP Clonase(TM)"
/note="recombination site for the Gateway(R) BP reaction"
CDS 3216..3257
/codon_start=1
/product="epitope tag from simian virus 5"
/label=V5 tag
/translation="GKPIPNPLLGLDST"
primer_bind complement(3712..3729)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 3718..3942
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(4075..4094)
/label=F1ori-R
/note="F1 origin, reverse primer"
protein_bind 4226..4273
/label=FRT
/bound_moiety="FLP recombinase from the Saccharomyces
cerevisiae 2u plasmid"
/note="FLP-mediated recombination occurs in the 8-bp core
sequence TCTAGAAA (Turan and Bode, 2011)."
CDS 4281..5303
/codon_start=1
/gene="aph(4)-Ia"
/product="aminoglycoside phosphotransferase from E. coli"
/label=HygR
/note="confers resistance to hygromycin"
/translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
"
polyA_signal 5433..5554
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(5470..5489)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 5524..5543
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(5603..5619)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5603..5619)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5616..5638)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5627..5643
/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(5651..5681)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5696..5717
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5834..5851)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6005..6593)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6085..6104)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6764..7624)
/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 7387..7406
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7625..7729)
/gene="bla"
/label=AmpR promoter
primer_bind complement(7804..7823)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata
1201 cgactcacta tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac
1261 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gcaggctcca
1321 ccatgtcgga cacgtggagc tctatccagg cccacaagaa gcagctggac tctctgcggg
1381 agaggctgca gcggaggcgg aagcaggact cggggcactt ggatctacgg aatccagagg
1441 cagcattgtc tccaaccttc cgtagtgaca gcccagtgcc tactgcaccc acctctggtg
1501 gccctaagcc cagcacagct tcagcagttc ctgaattagc tacagatcct gagttagaga
1561 agaagttgct acaccacctc tctgatctgg ccttaacatt gcccactgat gctgtgtcca
1621 tctgtcttgc catctccacg ccagatgctc ctgccactca agatggggta gaaagcctcc
1681 tgcagaagtt tgcagctcag gagttgattg aggtaaagcg aggtctccta caagatgatg
1741 cacatcctac tcttgtaacc tatgctgacc attccaagct ctctgccatg atgggtgctg
1801 tggcagaaaa gaagggccct ggggaggtag cagggactgt cacagggcag aagcggcgtg
1861 cagaacagga ctcgactaca gtagctgcct ttgccagttc gttagtctct ggtctgaact
1921 cttcagcatc ggaaccagca aaggagccag ccaagaaatc aaggaaacat gctgcctcag
1981 atgttgatct ggagatagag agccttctga accaacagtc cactaaggaa caacagagca
2041 agaaggtcag tcaggagatc ctagagctat taaatactac aacagccaag gaacaatcca
2101 ttgttgaaaa atttcgctct cgaggtcggg cccaagtgca agaattctgt gactatggaa
2161 ccaaggagga gtgcatgaaa gccagtgatg ctgatcgacc ctgtcgcaag ctgcacttca
2221 gacgaattat caataaacac actgatgagt ctttaggtga ctgctctttc cttaatacat
2281 gtttccacat ggatacctgc aagtatgttc aatatgaaat tgatgcttgc atggattctg
2341 aggcccctgg cagcaaagac cacacgccaa gccaggagct tgctcttaca cagagtgtcg
2401 gaggtgattc cagtgcagac cgactcttcc cacctcagtg gatctgttgt gatatccgct
2461 acctggacgt cagtatcttg ggcaagtttg cagttgtgat ggctgaccca ccctgggata
2521 ttcacatgga actgccctat gggaccctga cagatgatga gatgcgcagg ctcaacatac
2581 ccgtactaca ggatgatggc tttctcttcc tctgggtcac aggcagggcc atggagttgg
2641 ggagagaatg tctaaacctc tgggggtatg aacgggtaga tgaaattatt tgggtgaaga
2701 caaatcaact gcaacgcatc attcggacag gccgtacagg tcactggttg aaccatggga
2761 aggaacactg cttggttggt gtcaaaggaa atccccaagg cttcaaccag ggtctggatt
2821 gtgatgtgat cgtagctgag gttcgttcca ccagtcataa accagatgaa atctatggca
2881 tgattgaaag actatctcct ggcactcgca agattgagtt atttggacga ccacacaatg
2941 tgcaacccaa ctggatcacc cttggaaacc aactggatgg gatccaccta ctagacccag
3001 atgtggttgc acggttcaag caaaagtacc cagatggtat catctctaaa cctaagaatt
3061 tattggaccc agctttcttg tacaaagtgg ttgatatcca gcacagtggc ggccgctcga
3121 gtctagaggg cccgcggttc gaaaccagcg gaggcggcgg aagcggggga ggaggcagcg
3181 gcggaggcgg cagcgggggc ggcggaagcg ctaccggtaa gcctatccct aaccctctcc
3241 tcggtctcga ttctacgcgt accggtatgg cttctaactt tactcagttc gttctcgtcg
3301 acaatggcgg aactggcgac gtgactgtcg ccccaagcaa cttcgctaac ggggtcgctg
3361 aatggatcag ctctaactcg cgatcacagg cttacaaagt aacctgtagc gttcgtcaga
3421 gctctgcgca gaatcgcaaa tacaccatca aagtcgaggt gcctaaagtg gcaacccaga
3481 ctgttggtgg tgaagagctt cctgtagccg gatggagatc ttacttaaat atggaactaa
3541 ccattccaat tttcgccacg aattccgact gcgagcttat tgttaaggca atgcaaggtc
3601 tcctaaaaga tggaaacccg attccctcgg ccatcgcggc caactccggc atctacggag
3661 gtggaggtgg atctcgtacg ggttagtaat gagtttaaac ccgctgatca gcctcgactg
3721 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
3781 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
3841 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
3901 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
3961 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
4021 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
4081 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
4141 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
4201 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa
4261 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc
4321 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc
4381 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg
4441 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc
4501 cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc tcccgccgtg
4561 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg
4621 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc
4681 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg
4741 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg
4801 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg
4861 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca
4921 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct
4981 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg
5041 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct
5101 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg
5161 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg
5221 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca
5281 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct
5341 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
5401 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
5461 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
5521 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
5581 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
5641 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
5701 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
5761 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
5821 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
5881 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
5941 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
6001 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
6061 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
6121 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
6181 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
6241 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
6301 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
6361 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
6421 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
6481 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
6541 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
6601 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
6661 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
6721 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
6781 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
6841 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
6901 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
6961 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
7021 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
7081 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
7141 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
7201 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
7261 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
7321 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
7381 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
7441 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
7501 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
7561 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
7621 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
7681 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
7741 gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat ggtgcactct
7801 cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt
7861 ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga
7921 caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc
7981 cagatatacg cgttgacatt gattattgac taggcttttg caaaaagctt tgcaaagatg
8041 gataaagttt taaacagaga ggaatctttg cagctaatgg accttctagg tcttgaaagg
8101 agtgggaatt
//
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