| 质粒名称: | CoV2-M-IRES-E |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 177938 |
| 存储实验室: | Jennifer Doudna |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3.1 |
| 载体出品公司: | -- |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | -- |
| 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: | SARS-CoV-2 Membrane and Envelope proteins (original Wuhan Hu1 sequence) |
| 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: | Gibson克隆 |
| 5’克隆位点: | -- |
| 3’克隆位点: | -- |
| 5’测序引物 | -- |
| 3’测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | CoV2-M-IRES-E was a gift from Jennifer Doudna (Addgene plasmid # 177938 ; http://n2t.net/addgene:177938 ; RRID:Addgene_177938) |
| 参考文献部分: | Rapid assessment of SARS-CoV-2 evolved variants using virus-like particles. Syed AM, Taha TY, Tabata T, Chen IP, Ciling A, Khalid MM, Sreekumar B, Chen PY, Hayashi JM, Soczek KM, Ott M, Doudna JA. Science. 2021 Nov 4:eabl6184. doi: 10.1126/science.abl6184. 10.1126/science.abl6184 PubMed 34735219 |
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 CoV2-M-IRES-E 7521 bp ds-DNA circular SYN 16-NOV-2021
DEFINITION Transient mammalian expression of codon optimized SARS-CoV-2
Membrane and Envelope proteins (original Wuhan Hu1 sequence). Used
for generating RNA packaging virus-like particles..
ACCESSION .
VERSION .
KEYWORDS CoV2-M-IRES-E
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7521)
AUTHORS Syed AM, Taha TY, Tabata T, Chen IP, Ciling A, Khalid MM, Sreekumar
B, Chen PY, Hayashi JM, Soczek KM, Ott M, Doudna JA
TITLE Rapid assessment of SARS-CoV-2 evolved variants using virus-like
particles.
JOURNAL Science. 2021 Nov 4:eabl6184. doi: 10.1126/science.abl6184.
PUBMED 34735219
REFERENCE 2 (bases 1 to 7521)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Nov 16, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7521
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 91..553
/label=IRES
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
primer_bind complement(147..164)
/label=IRES reverse
/note="IRES internal ribosome entry site, reverse primer.
Also called pCDH-rev"
primer_bind 374..393
/label=IRES-F
/note="IRES internal ribosome entry site, forward primer"
misc_feature 837..1425
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(890..910)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(1308..1319)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(1515..1532)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 1521..1745
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 1791..2219
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1878..1897)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2088..2109
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(2228..2248)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 2233..2562
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 2413..2548
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2475..2494
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2629..3423
/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(2683..2702)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 3293..3312
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 3597..3718
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3634..3653)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3688..3707
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(3767..3783)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3767..3783)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(3780..3802)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 3791..3807
/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(3815..3845)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 3860..3881
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(3998..4015)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(4169..4757)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(4249..4268)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4928..5788)
/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 5551..5570
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(5789..5893)
/gene="bla"
/label=AmpR promoter
primer_bind complement(5968..5987)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 6159..6538
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 6539..6742
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 6693..6713
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 6787..6806
/label=T7
/note="T7 promoter, forward primer"
promoter 6787..6805
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 6825..6834
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
ORIGIN
1 taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt
61 ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt
121 gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt
181 cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct
241 ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt
301 ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt
361 ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa
421 ggtaccccat tgtatgggat ctgatctggg gcctcggtgc acatgcttta catgtgttta
481 gtcgaggtta aaaaaacgtc taggcccccc gaaccacggg gacgtggttt tcctttgaaa
541 aacacgatga taagcttgcc acaacccaca aggagaaatt cgccgccacc atgtacagct
601 tcgtatcaga agaaaccggg acactgatcg taaattctgt gctcttgttt ctggcattcg
661 tcgtatttct cctcgtcaca ctggcaattc tgactgcatt gaggctttgc gcctactgtt
721 gtaacattgt caatgtatct ctcgtgaaac cctcattcta cgtttacagc agggtgaaga
781 atctcaattc tagcagggtg ccggatctcc tcgttcttta actcgaccga ctcgacaatc
841 aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt
901 ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg
961 ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc
1021 ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt
1081 ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg
1141 ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg
1201 gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct
1261 gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc
1321 cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg catcttcgcc
1381 ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctgga attaattcga
1441 gctcggtacc tttaagacca atgacttaca aggcagcctc gagtctagag ggcccgttta
1501 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
1561 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
1621 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
1681 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
1741 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
1801 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
1861 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
1921 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
1981 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
2041 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
2101 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
2161 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
2221 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
2281 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
2341 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
2401 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
2461 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
2521 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
2581 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
2641 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
2701 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
2761 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
2821 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
2881 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
2941 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
3001 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
3061 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
3121 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
3181 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
3241 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
3301 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
3361 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
3421 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
3481 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
3541 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
3601 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
3661 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
3721 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
3781 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
3841 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
3901 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
3961 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
4021 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
4081 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
4141 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
4201 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
4261 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
4321 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
4381 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
4441 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
4501 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
4561 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
4621 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
4681 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
4741 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
4801 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
4861 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
4921 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
4981 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
5041 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
5101 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
5161 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
5221 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
5281 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
5341 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
5401 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
5461 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
5521 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
5581 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
5641 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
5701 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
5761 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
5821 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
5881 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
5941 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
6001 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
6061 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
6121 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
6181 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
6241 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
6301 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
6361 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
6421 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
6481 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
6541 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
6601 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
6661 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
6721 ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt actggcttat
6781 cgaaattaat acgactcact atagggagac ccaagctggc tagcgccacc atggccgact
6841 caaatgggac cattactgta gaggagctga aaaaactgtt ggagcaatgg aaccttgtga
6901 ttggatttct cttcttgaca tggatttgtc tcctgcaatt cgcatacgcg aatcgcaacc
6961 gattcctcta tattataaaa cttatattcc tttggctcct gtggcctgtg acccttgcct
7021 gcttcgtact cgcggcggtg tatcgaataa actggataac tggcggcata gcaatagcta
7081 tggcatgcct ggtaggtttg atgtggctct cctattttat agcgtcattc agactttttg
7141 cacgcacgcg aagcatgtgg tcatttaacc ctgaaacaaa tatcttgctt aatgtccccc
7201 tgcatggcac aattttgacc cgaccgctcc ttgaatcaga attggtgatt ggagccgtga
7261 tcctgagggg ccacttgagg atcgcaggac atcatcttgg tcgctgtgat attaaggatt
7321 tgccaaaaga aataactgta gctaccagtc gaactttgag ctattacaag cttggagcgt
7381 cccaaagggt cgctggcgat agtggatttg cagcctattc acgctacagg atagggaact
7441 ataaactgaa tacagaccac tcatcatctt ctgataacat tgctttgctc gtccagtaag
7501 gatcccgccc ctctccctcc c
//
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