| 质粒名称: | SARS-CoV-2 Nucleocapsid-3xHA-IRES-mcherry |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 173030 |
| 存储实验室: | Mohamed Fareh |
|
载体骨架基本信息 |
|
| 载体名称: | MscV-IRES-mcherry |
| 载体出品公司: | -- |
| 空载体大小: | 6502 bp |
| 完整质粒大小: | 7759 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳动物表达，慢病毒 |
| 筛选标记: | mcherry |
| 载体抗性: | 氨苄青霉素，100μg/mL |
| 生长温度: | 37 ℃ |
| 宿主菌: | NEB Stable |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | SARS-CoV-2 Nucleocapsid phosphoprotein |
| 物种来源: | SARS冠状病毒2型 |
| 插入大小: | 1257 bp |
| 基因突变: | -- |
| GenBank编号 | BCG67561.1 |
| 启动子: | Gag（截断） |
| 标签/融合蛋白: | 3xHA（插入件上的C端子）IRES mcherry（主干上的C终端） |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | BamHI(未破坏) |
| 3’克隆位点: | BamHI(未破坏) |
| 5’测序引物 | ATGTCTGATAATGGACCCCAAAATCAGC |
| 3’测序引物 | GGCCTGAGTTGAGTCAGCACT |
|
文献引用方法 |
|
| 材料和方法部分: | SARS-CoV-2 Nucleocapsid-3xHA-IRES-mcherry was a gift from Mohamed Fareh (Addgene plasmid # 173030 ; http://n2t.net/addgene:173030 ; RRID:Addgene_17303 |
| 参考文献部分: | Reprogrammed CRISPR-Cas13b suppresses SARS-CoV-2 replication and circumvents its mutational escape through mismatch tolerance. Fareh M, Zhao W, Hu W, Casan JML, Kumar A, Symons J, Zerbato JM, Fong D, Voskoboinik I, Ekert PG, Rudraraju R, Purcell DFJ, Lewin SR, Trapani JA. Nat Commun. 2021 Jul 13;12(1):4270. doi: 10.1038/s41467-021-24577-9. 10.1038/s41467-021-24577-9 PubMed 342573110) |
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 SARS-CoV-2 Nucleocapsid-3xHA-IRES-mcherry 7867 bp ds-DNA circular SYN 31-AUG-2021
DEFINITION To express SARS-CoV-2 Nucleocapsid protein tagged with 3xHA and in
frame with mcherry (NCP-IRES-mcherry) in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS SARS-CoV-2 Nucleocapsid-3xHA-IRES-mcherry
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7867)
AUTHORS Fareh M, Zhao W, Hu W, Casan JML, Kumar A, Symons J, Voskoboinik I,
Ekert PG, Rudraraju R, Lewin SR, Trapani JA
TITLE Reprogrammed CRISPR-Cas13b suppresses SARS-CoV-2 replication and
circumvents its mutational escape through mismatch tolerance
JOURNAL BioRxiv 2020
REFERENCE 2 (bases 1 to 7867)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 31, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7867
/organism="synthetic DNA construct"
/mol_type="other DNA"
misc_feature 170..511
/label=MESV Psi
/note="packaging signal of murine embryonic stem cell
virus"
CDS 578..994
/label=gag (truncated)
/note="truncated Moloney murine leukemia virus (MMLV) gag
gene lacking the start codon"
primer_bind 924..946
/label=pLXSN 5'
/note="Murine stem cell virus, forward primer. Also called
MSCV"
primer_bind 962..978
/label=pBABE 5'
/note="Psi packaging signal, forward primer for pBABE
vectors"
CDS 2282..2362
/codon_start=1
/product="three tandem HA epitope tags"
/label=3xHA
/translation="YPYDVPDYAYPYDVPDYAYPYDVPDYA"
primer_bind complement(2571..2588)
/label=IRES reverse
/note="IRES internal ribosome entry site, reverse primer.
Also called pCDH-rev"
primer_bind 2798..2817
/label=IRES-F
/note="IRES internal ribosome entry site, forward primer"
primer_bind complement(3124..3142)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 3392..3411
/label=mCherry-F
/note="mCherry, forward primer"
LTR 3851..4365
/label=3' LTR
/note="3' long terminal repeat from murine embryonic stem
cell virus"
protein_bind 4527..4543
/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(4551..4581)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4596..4617
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(4734..4751)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(4905..5493)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(4985..5004)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5664..6524)
/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 6287..6306
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6525..6629)
/gene="bla"
/label=AmpR promoter
primer_bind 6697..6715
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(6753..6775)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 6875..6894
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 7088..7110
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 7238..7257
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
LTR 7457..106
/label=5' LTR
/note="5' long terminal repeat from murine embryonic stem
cell virus"
ORIGIN
1 tattcccaat aaagcctctt gctgtttgca tccgaatcgt ggactcgctg atccttggga
61 gggtctcctc agattgattg actgcccacc tcgggggtct ttcatttgga ggttccaccg
121 agatttggag acccctgccc agggaccacc gacccccccg ccgggaggta agctggccag
181 cggtcgtttc gtgtctgtct ctgtctttgt gcgtgtttgt gccggcatct aatgtttgcg
241 cctgcgtctg tactagttag ctaactagct ctgtatctgg cggacccgtg gtggaactga
301 cgagttctga acacccggcc gcaaccctgg gagacgtccc agggactttg ggggccgttt
361 ttgtggcccg acctgaggaa gggagtcgat gtggaatccg accccgtcag gatatgtggt
421 tctggtagga gacgagaacc taaaacagtt cccgcctccg tctgaatttt tgctttcggt
481 ttggaaccga agccgcgcgt cttgtctgct gcagcgctgc agcatcgttc tgtgttgtct
541 ctgtctgact gtgtttctgt atttgtctga aaattagggc cagactgtta ccactccctt
601 aagtttgacc ttaggtcact ggaaagatgt cgagcggatc gctcacaacc agtcggtaga
661 tgtcaagaag agacgttggg ttaccttctg ctctgcagaa tggccaacct ttaacgtcgg
721 atggccgcga gacggcacct ttaaccgaga cctcatcacc caggttaaga tcaaggtctt
781 ttcacctggc ccgcatggac acccagacca ggtcccctac atcgtgacct gggaagcctt
841 ggcttttgac ccccctccct gggtcaagcc ctttgtacac cctaagcctc cgcctcctct
901 tcctccatcc gccccgtctc tcccccttga acctcctcgt tcgaccccgc ctcgatcctc
961 cctttatcca gccctcactc cttctctagg cgccggaatt cgtttaaacg gatccatgtc
1021 tgataatgga ccccaaaatc agcgaaatgc accccgcatt acgtttggtg gaccctcaga
1081 ttcaactggc agtaaccaga atggagaacg cagtggggcg cgatcaaaac aacgtcggcc
1141 ccaaggttta cccaataata ctgcgtcttg gttcaccgct ctcactcaac atggcaagga
1201 agaccttaaa ttccctcgag gacaaggcgt tccaattaac accaatagca gtccagatga
1261 ccaaattggc tactaccgaa gagctaccag acgaattcgt ggtggtgacg gtaaaatgaa
1321 agatctcagt ccaagatggt atttctacta cctaggaact gggccagaag ctggacttcc
1381 ctatggtgct aacaaagacg gcatcatatg ggttgcaact gagggagcct tgaatacacc
1441 aaaagatcac attggcaccc gcaatcctgc taacaatgct gcaatcgtgc tacaacttcc
1501 tcaaggaaca acattgccaa aaggcttcta cgcagaaggg agcagaggcg gcagtcaagc
1561 ctcttctcgt tcctcatcac gtagtcgcaa cagttcaaga aattcaactc caggcagcag
1621 taggggaact tctcctgcta gaatggctgg caatggcggt gatgctgctc ttgctttgct
1681 gctgcttgac agattgaacc agcttgagag caaaatgtct ggtaaaggcc aacaacaaca
1741 aggccaaact gtcactaaga aatctgctgc tgaggcttct aagaagcctc ggcaaaaacg
1801 tactgccact aaagcataca atgtaacaca agctttcggc agacgtggtc cagaacaaac
1861 ccaaggaaat tttggggacc aggaactaat cagacaagga actgattaca aacattggcc
1921 gcaaattgca caatttgccc ccagcgcttc agcgttcttc ggaatgtcgc gcattggcat
1981 ggaagtcaca ccttcgggaa cgtggttgac ctacacaggt gccatcaaat tggatgacaa
2041 agatccaaat ttcaaagatc aagtcatttt gctgaataag catattgacg catacaaaac
2101 attcccacca acagagccta aaaaggacaa aaagaagaag gctgatgaaa ctcaagcctt
2161 accgcagaga cagaagaaac agcaaactgt gactcttctt cctgctgcag atttggatga
2221 tttctccaaa caattgcaac aatccatgag cagtgctgac tcaactcagg ccggtagttc
2281 ctacccatac gatgttccag attacgctta tccctacgac gtgcctgatt atgcataccc
2341 atatgatgtc cccgactatg cctaaggatc cagatctcca gcacagtgga cgcgtctcga
2401 gcgggatcaa ttccgccccc cccctaacgt tactggccga agccgcttgg aataaggccg
2461 gtgtgcgttt gtctatatgt tattttccac catattgccg tcttttggca atgtgagggc
2521 ccggaaacct ggccctgtct tcttgacgag cattcctagg ggtctttccc ctctcgccaa
2581 aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag
2641 acaaacaacg tctgtagcga ccctttgcag gcagcggaac cccccacctg gcgacaggtg
2701 cctctgcggc caaaagccac gtgtataaga tacacctgca aaggcggcac aaccccagtg
2761 ccacgttgtg agttggatag ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa
2821 caaggggctg aaggatgccc agaaggtacc ccattgtatg ggatctgatc tggggcctcg
2881 gtgcacatgc tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca
2941 cggggacgtg gttttccttt gaaaaacacg ataataccat ggtgagcatg gacgaggagg
3001 ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga
3061 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
3121 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc
3181 ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact
3241 tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
3301 gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga
3361 agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct
3421 gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc
3481 agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg
3541 ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct
3601 cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca
3661 ccggcggcat ggacgagctg tacaagtaaa gcggccgcga ctctagagtc gacctgcagg
3721 catgcaagct tcaggtagcc ggctaacgtt aacaaccggt acctctagaa ctatagctag
3781 catgcgcaaa tttaaagcgc tgatatcgat aaaataaaag attttattta gtctccagaa
3841 aaagggggga atgaaagacc ccacctgtag gtttggcaag ctagcttaag taacgccatt
3901 ttgcaaggca tggaaaatac ataactgaga atagagaagt tcagatcaag gttaggaaca
3961 gagagacagc agaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct
4021 cagggccaag aacagatggt ccccagatgc ggtcccgccc tcagcagttt ctagagaacc
4081 atcagatgtt tccagggtgc cccaaggacc tgaaatgacc ctgtgcctta tttgaactaa
4141 ccaatcagtt cgcttctcgc ttctgttcgc gcgcttctgc tccccgagct caataaaaga
4201 gcccacaacc cctcactcgg cgcgccagtc ctccgataga ctgcgtcgcc cgggtacccg
4261 tgtatccaat aaaccctctt gcagttgcat ccgacttgtg gtctcgctgt tccttgggag
4321 ggtctcctct gagtgattga ctacccgtca gcgggggtct ttcatgggta acagtttctt
4381 gaagttggag aacaacattc tgagggtagg agtcgaatat taagtaatcc tgactcaatt
4441 agccactgtt ttgaatccac atactccaat actcctgaaa tagttcatta tggacagcgc
4501 agaaagagct ggggagaatt gtgaaattgt tatccgctca caattccaca caacatacga
4561 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
4621 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
4681 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
4741 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
4801 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
4861 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
4921 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
4981 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
5041 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
5101 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
5161 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
5221 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
5281 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
5341 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
5401 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
5461 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
5521 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
5581 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
5641 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
5701 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
5761 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
5821 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
5881 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
5941 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
6001 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
6061 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
6121 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
6181 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
6241 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
6301 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
6361 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
6421 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
6481 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
6541 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
6601 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
6661 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
6721 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg
6781 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg
6841 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga
6901 gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg
6961 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
7021 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
7081 gggttttccc agtcacgacg ttgtaaaacg acggcgcaag gaatggtgca tgcaaggaga
7141 tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc
7201 tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc
7261 cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggcgat
7321 tagtccaatt tgttaaagac aggatatcag tggtccaggc tctagttttg actcaacaat
7381 atcaccagct gaagcctata gagtacgagc catagataaa ataaaagatt ttatttagtc
7441 tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta gcttaagtaa
7501 cgccattttg caaggcatgg aaaatacata actgagaata gagaagttca gatcaaggtt
7561 aggaacagag agacagcaga atatgggcca aacaggatat ctgtggtaag cagttcctgc
7621 cccggctcag ggccaagaac agatggtccc cagatgcggt cccgccctca gcagtttcta
7681 gagaaccatc agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt
7741 gaactaacca atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa
7801 taaaagagcc cacaacccct cactcggcgc gccagtcctc cgatagactg cgtcgcccgg
7861 gtacccg
//
