| 质粒名称: | pcDNA3-sACE2(WT)-8his |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 149268 |
| 存储实验室: | Erik Procko |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3.1(+) |
| 载体出品公司: | Invitrogen |
| 空载体大小: | 5332 bp |
| 完整质粒大小: | 7234 bp |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 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: | Angiotensin-converting enzyme 2 / ACE2 / peptidyl-dipeptidase A |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 1902 bp |
| 基因突变: | Extracellular, soluble protease domain (a.a. M1-D615) |
| GenBank编号 | NM_021804.1 |
| 启动子: | CMV |
| 标签/融合蛋白: | Short GSG linker and 8his tag (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | NheI (not destroyed) |
| 3’克隆位点: | XhoI (not destroyed) |
| 5’测序引物 | CMV-F |
| 3’测序引物 | BGH-rev |
|
文献引用方法 |
|
| 材料和方法部分: |
pcDNA3-sACE2(WT)-8his was a gift from Erik Procko (Addgene plasmid # 149268 ; http://n2t.net/addgene:149268 ; RRID:Addgene_149268)
|
| 参考文献部分: | Engineering human ACE2 to optimize binding to the spike protein of SARS coronavirus 2. Chan KK, Dorosky D, Sharma P, Abbasi SA, Dye JM, Kranz DM, Herbert AS, Procko E. Science. 2020 Sep 4;369(6508):1261-1265. doi: 10.1126/science.abc0870. Epub 2020 Aug 4. 10.1126/science.abc0870 PubMed 32753553 |
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
-8his质粒图谱.png)
载体简介
载体序列
LOCUS pcDNA3-sACE2(WT)-8his 7237 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Mammalian expression plasmid for his-tagged sACE2.
ACCESSION .
VERSION .
KEYWORDS pcDNA3-sACE2(WT)-8his
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7237)
AUTHORS Chan KK, Dorosky D, Sharma P, Abbasi SA, Dye JM, Kranz DM, Herbert
AS, Procko E
TITLE Engineering human ACE2 to optimize binding to the spike protein of
SARS coronavirus 2.
JOURNAL Science. 2020 Sep 4;369(6508):1261-1265. doi:
10.1126/science.abc0870. Epub 2020 Aug 4.
PUBMED 32753553
REFERENCE 2 (bases 1 to 7237)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7237
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(32..49)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 38..262
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 308..736
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(395..414)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 605..626
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(745..765)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 750..1079
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 930..1065
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 992..1011
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 1146..1940
/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(1200..1219)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 1810..1829
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 2114..2235
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2151..2170)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 2205..2224
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(2284..2300)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(2284..2300)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2297..2319)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 2308..2324
/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(2332..2362)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2377..2398
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(2515..2532)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2686..3274)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2766..2785)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3445..4305)
/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 4068..4087
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(4306..4410)
/gene="bla"
/label=AmpR promoter
primer_bind complement(4485..4504)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 4676..5055
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 5056..5259
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 5210..5230
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 5304..5323
/label=T7
/note="T7 promoter, forward primer"
promoter 5304..5322
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 7202..7225
/codon_start=1
/product="8xHis affinity tag"
/label=8xHis
/translation="HHHHHHHH"
ORIGIN
1 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
61 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
121 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
181 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
241 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
301 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
361 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
421 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
481 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
541 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
601 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
661 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
721 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
781 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
841 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
901 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
961 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
1021 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
1081 agctcccggg agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt
1141 ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
1201 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
1261 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
1321 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
1381 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
1441 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
1501 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
1561 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
1621 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
1681 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
1741 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
1801 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
1861 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
1921 ttcttgacga gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc
1981 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
2041 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
2101 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
2161 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
2221 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
2281 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
2341 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
2401 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
2461 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
2521 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
2581 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
2641 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
2701 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
2761 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
2821 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
2881 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
2941 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
3001 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
3061 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
3121 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
3181 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
3241 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
3301 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
3361 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
3421 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
3481 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
3541 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
3601 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
3661 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
3721 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
3781 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
3841 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
3901 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
3961 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
4021 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
4081 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
4141 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
4201 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
4261 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
4321 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
4381 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
4441 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
4501 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
4561 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
4621 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
4681 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
4741 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
4801 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
4861 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
4921 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
4981 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
5041 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
5101 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
5161 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
5221 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
5281 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
5341 cgccaccatg tcaagctctt cctggctcct tctcagcctt gttgctgtaa ctgctgctca
5401 gtccaccatt gaggaacagg ccaagacatt tttggacaag tttaaccacg aagccgaaga
5461 cctgttctat caaagttcac ttgcttcttg gaattataac accaatatta ctgaagagaa
5521 tgtccaaaac atgaataatg ctggggacaa atggtctgcc tttttaaagg aacagtccac
5581 acttgcccaa atgtatccac tacaagaaat tcagaatctc acagtcaagc ttcagctgca
5641 ggctcttcag caaaatgggt cttcagtgct ctcagaagac aagagcaaac ggttgaacac
5701 aattctaaat acaatgagca ccatctacag tactggaaaa gtttgtaacc cagataatcc
5761 acaagaatgc ttattacttg aaccaggttt gaatgaaata atggcaaaca gtttagacta
5821 caatgagagg ctctgggctt gggaaagctg gagatctgag gtcggcaagc agctgaggcc
5881 attatatgaa gagtatgtgg tcttgaaaaa tgagatggca agagcaaatc attatgagga
5941 ctatggggat tattggagag gagactatga agtaaatggg gtagatggct atgactacag
6001 ccgcggccag ttgattgaag atgtggaaca tacctttgaa gagattaaac cattatatga
6061 acatcttcat gcctatgtga gggcaaagtt gatgaatgcc tatccttcct atatcagtcc
6121 aattggatgc ctccctgctc atttgcttgg tgatatgtgg ggtagatttt ggacaaatct
6181 gtactctttg acagttccct ttggacagaa accaaacata gatgttactg atgcaatggt
6241 ggaccaggcc tgggatgcac agagaatatt caaggaggcc gagaagttct ttgtatctgt
6301 tggtcttcct aatatgactc aaggattctg ggaaaattcc atgctaacgg acccaggaaa
6361 tgttcagaaa gcagtctgcc atcccacagc ttgggacctg gggaagggcg acttcaggat
6421 tcttatgtgc acaaaggtga caatggacga cttcctgaca gctcatcatg agatggggca
6481 tatccagtat gatatggcat atgctgcaca accttttctg ctaagaaatg gagctaatga
6541 aggattccat gaagctgttg gggaaatcat gtcactttct gcagccacac ctaagcattt
6601 aaaatccatt ggtcttctgt cacccgattt tcaagaagac aatgaaacag aaataaactt
6661 cctgctcaaa caagcactca cgattgttgg gactctgcca tttacttaca tgttagagaa
6721 gtggaggtgg atggtcttta aaggggaaat tcccaaagac cagtggatga aaaagtggtg
6781 ggagatgaag cgagagatag ttggggtggt ggaacctgtg ccccatgatg aaacatactg
6841 tgaccccgca tctctgttcc atgtttctaa tgattactca ttcattcgat attacacaag
6901 gaccctttac caattccagt ttcaagaagc actttgtcaa gcagctaaac atgaaggccc
6961 tctgcacaaa tgtgacatct caaactctac agaagctgga cagaaactgt tcaatatgct
7021 gaggcttgga aaatcagaac cctggaccct agcattggaa aatgttgtag gagcaaagaa
7081 catgaatgta aggccactgc tcaactactt tgagccctta tttacctggc tgaaagacca
7141 gaacaagaat tcttttgtgg gatggagtac cgactggagt ccatatgcag acggatccgg
7201 tcaccatcac caccaccatc atcattgata actcgag
//
