| 质粒名称: | pPolI_SARS-CoV-2_NLuc |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 183953 |
| 存储实验室: | AJ te Velthuis |
|
载体骨架基本信息 |
|
| 载体名称: | pMK-RQ |
| 载体出品公司: | Invitrogen |
| 空载体大小: | 2278 bp |
| 完整质粒大小: | 4830 bp |
| 载体类型: | 哺乳细胞表达 |
| 载体抗性: | Kanamycin, 50 μg/mL |
| 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 minigenome with Nano luciferase |
| 物种来源: | SARS coronavirus 2 |
| 插入大小: | 2252 bp |
| 基因突变: |
N (a.k.a. GU280_gp10)
Entrez GeneORF10 (a.k.a. GU280_gp11)
|
| GenBank编号 | NC_045512.2 |
| 启动子: | Mammalian Pol I |
| 标签/融合蛋白: | 人细胞色素c氧化酶亚基VIII（N端） |
| 克隆方法: | 未知 |
| 5’测序引物 | GTGAGCGCGACGTAATAC |
| 3’测序引物 | GCTGGCACGACAGGTTTC |
|
文献引用方法 |
|
| 材料和方法部分: | pPolI_SARS-CoV-2_NLuc was a gift from AJ te Velthuis (Addgene plasmid # 183953 ; http://n2t.net/addgene:183953 ; RRID:Addgene_183953) |
| 参考文献部分: | A SARS-CoV-2 mini-genome assay based on negative-sense RNA to study replication inhibitors and emerging mutations. Vial TK, Oade MS, Russell CA, Eggink D, te Velthuis AJW. bioRxiv 2021.06.28.450211 10.1101/2021.06.28.450211 |
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 AJ te Velthuis 4830 bp ds-DNA circular SYN 24-JUN-2022
DEFINITION Encodes SARS-CoV-2 minigenome with nano-luciferase reporter gene.
Minigenome is under control of the Pol I promoter.
ACCESSION .
VERSION .
KEYWORDS pPolI_SARS-CoV-2_NLuc
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4830)
AUTHORS Vial TK, Oade MS, Russell CA, Eggink D, te Velthuis AJW
TITLE A SARS-CoV-2 mini-genome assay based on negative-sense RNA to study
replication inhibitors and emerging mutations
JOURNAL bioRxiv 2021.06.28.450211
REFERENCE 2 (bases 1 to 4830)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jun 24, 2022 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4830
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 278..300
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 292..309
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 293..309
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 320..339
/label=T7
/note="T7 promoter, forward primer"
promoter 320..338
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 343..354
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
CDS complement(2232..2747)
/codon_start=1
/product="NanoLuc(R) luciferase"
/label=Nluc
/note="human codon-optimized"
/translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
RLCERILA"
primer_bind complement(3036..3053)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3146..3734)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3226..3245)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3908..4702)
/codon_start=1
/product="aminoglycoside phosphotransferase"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
promoter complement(4703..4807)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
121 gatagggttg agtggccgct acagggcgct cccattcgcc attcaggctg cgcaactgtt
181 gggaagggcg tttcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
241 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
301 acggccagtg agcgcgacgt aatacgactc actatagggc gaattgaagg aaggccgtca
361 aggccgcata agcttacggg ccggcccctg cttgtggcac gggcggccgg gagggcgtcc
421 ccggcccggc gctgctcccg cgtgtgtcct ggggttgacc agagggcccc gggcgctccg
481 tgtgtggctg cgatggtggc gtttttgggg acaggtgtcc gtgtcgcgcg tcgcctgggc
541 cggcggcgtg gtcggtgacg cgacctcccg gccccggggg aggtatatct ttcgctccga
601 gtcggcattt tgggccgccg ggttattttt tttttttgtc attctcctaa gaagctatta
661 aaatcacatg gggatagcac tactaaaatt aattttacac attagggctc ttccatatag
721 gcagctctcc ctagcattgt tcactgtaca ctcgatcgta ctccgcgtgg cctcggtgaa
781 aatgtggtgg ctctttcaag tcctccctaa tgttacacac tgattaaaga ttgctatgtg
841 agattaaagt taactacatc tacttgtgct atgtagttac gagaattcat tctgcacaag
901 agtagactat atatcgtaaa cggaaaagcg aaaacgttta tatagcccat ctgccttgtg
961 tggtctgcat gagtttaggc ctgagttgag tcagcactgc tcatggattg ttgcaattgt
1021 ttggagaaat catccaaatc tgcagcagga agaagagtca cagtttgctg tttcttctgt
1081 ctctgcggta aggcttgagt ttcatcagcc ttcttctttt tgtccttttt aggctctgtt
1141 ggtgggaatg ttttgtatgc gtcaatatgc ttattcagca aaatgacttg atctttgaaa
1201 tttggatctt tgtcatccaa tttgatggca cctgtgtagg tcaaccacgt tcccgaaggt
1261 gtgacttcca tgccaatgcg cgacattccg aagaacgctg aagcgctggg ggcaaattgt
1321 gcaatttgcg gccaatgttt gtaatcagtt ccttgtctga ttagttcctg gtccccaaaa
1381 tttccttggg tttgttctgg accacgtctg ccgaaagctt gtgttacatt gtatgcttta
1441 gtggcagtac gtttttgccg aggcttctta gaagcctcag cagcagattt cttagtgaca
1501 gtttggcctt gttgttgttg gcctttacca gacattttgc tctcaagctg gttcaatctg
1561 tcaagcagca gcaaagcaag agcagcatca ccgccattgc cagccattct agcaggagaa
1621 gttcccctac tgctgcctgg agttgaattt cttgaactgt tgcgactacg tgatgaggaa
1681 cgagaagagg cttgactgcc gcctctgctc ccttctgcgt agaagccttt tggcaatgtt
1741 gttccttgag gaagttgtag cacgattgca gcattgttag caggattgcg ggtgccaatg
1801 tgatcttttg gtgtattcaa ggctccctca gttgcaaccc atatgatgcc gtctttgtta
1861 gcaccatagg gaagtccagc ttctggccca gttcctaggt agtagaaata ccatcttgga
1921 ctgagatctt tcattttacc gtcaccacca cgaattcgtc tggtagctct tcggtagtag
1981 ccaatttggt catctggact gctattggtg ttaattggaa cgccttgtcc tcgagggaat
2041 ttaaggtctt ccttgccatg ttgagtgaga gcggtgaacc aagacgcagt attattgggt
2101 aaaccttggg gccgacgttg ttttgatcgc gccccactgc gttctccatt ctggttactg
2161 ccagttgaat ctgagggtcc accaaacgta atgcggggtg catttcgctg attttggggt
2221 ccattatcag attacgccag aatgcgttcg cacagccgcc agccggtcac tccgttgatg
2281 gttactcgga acagcaggga gccgtcgggg ttgatcaggc gctcgtcgat aattttgttg
2341 ccgttccaca gggtccctgt tacagtgatc tttttgccgt cgaacacggc gatgccttca
2401 tacggccgtc cgaaatagtc gatcatgttc ggcgtaaccc cgtcgattac cagtgtgcca
2461 tagtgcagga tcaccttaaa gtgatgatca tccacagggt acaccacctt aaaaattttt
2521 tcgatctggc ccatttggtc gccgctcaga ccttcatacg ggatgatgac atggatgtcg
2581 atcttcagcc cattttcacc gctcaggaca atcctttgga tcggagttac ggacaccccg
2641 agattctgaa acaaactgga cacacctccc tgttcaagga cttggtccag gttgtagccg
2701 gctgtctgtc gccagtcccc aacgaaatct tcgagtgtga agaccatttt agtttgttcg
2761 tttagagaac agatctacaa gagatcgaaa gttggttggt ttgttacctg ggaaggtata
2821 aacctttaat ggccggcatg gtcccagcct cctcgctggc gccggctggg caacattccg
2881 aggggaccgt cccctgggta atggcgaatg ggacctctag actgggcctc atgggccttc
2941 ctttcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta acatggtcat
3001 agctgtttcc ttgcgtattg ggcgctctcc gcttcctcgc tcactgactc gctgcgctcg
3061 gtcgttcggg taaagcctgg ggtgcctaat gagcaaaagg ccagcaaaag gccaggaacc
3121 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
3181 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
3241 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
3301 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
3361 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
3421 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
3481 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
3541 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
3601 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
3661 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
3721 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
3781 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
3841 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
3901 acagttatta gaaaaattca tccagcagac gataaaacgc aatacgctgg ctatccggtg
3961 ccgcaatgcc atacagcacc agaaaacgat ccgcccattc gccgcccagt tcttccgcaa
4021 tatcacgggt ggccagcgca atatcctgat aacgatccgc cacgcccaga cggccgcaat
4081 caataaagcc gctaaaacgg ccattttcca ccataatgtt cggcaggcac gcatcaccat
4141 gggtcaccac cagatcttcg ccatccggca tgctcgcttt cagacgcgca aacagctctg
4201 ccggtgccag gccctgatgt tcttcatcca gatcatcctg atccaccagg cccgcttcca
4261 tacgggtacg cgcacgttca atacgatgtt tcgcctgatg atcaaacgga caggtcgccg
4321 ggtccagggt atgcagacga cgcatggcat ccgccataat gctcactttt tctgccggcg
4381 ccagatggct agacagcaga tcctgacccg gcacttcgcc cagcagcagc caatcacggc
4441 ccgcttcggt caccacatcc agcaccgccg cacacggaac accggtggtg gccagccagc
4501 tcagacgcgc cgcttcatcc tgcagctcgt tcagcgcacc gctcagatcg gttttcacaa
4561 acagcaccgg acgaccctgc gcgctcagac gaaacaccgc cgcatcagag cagccaatgg
4621 tctgctgcgc ccaatcatag ccaaacagac gttccaccca cgctgccggg ctacccgcat
4681 gcaggccatc ctgttcaatc atactcttcc tttttcaata ttattgaagc atttatcagg
4741 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
4801 ttccgcgcac atttccccga aaagtgccac
//
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