| 质粒名称: | pRK5-HA-Ubiquitin-K27R |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 121155 |
| 存储实验室: | Josef Kittler |
|
载体骨架基本信息 |
|
| 载体名称: | pRK5-HA |
| 载体出品公司: | Ted Dawson, Addgene plasmid #17608 |
| 完整质粒大小: | 4996 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: | Ubiquitin C / Ub |
| 物种来源: | H. sapiens (human) |
| 基因突变: | K27 mutated to an R |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | unknown (unknown if destroyed) |
| 3’克隆位点: | unknown (unknown if destroyed) |
| 5’测序引物 | 未知 |
|
文献引用方法 |
|
| 材料和方法部分: |
pRK5-HA-Ubiquitin-K27R was a gift from Josef Kittler (Addgene plasmid # 121155 ; http://n2t.net/addgene:121155 ; RRID:Addgene_121155)
|
| 参考文献部分: | Lysine 27 ubiquitination of the mitochondrial transport protein Miro is dependent on serine 65 of the Parkin ubiquitin ligase. Birsa N, Norkett R, Wauer T, Mevissen TE, Wu HC, Foltynie T, Bhatia K, Hirst WD, Komander D, Plun-Favreau H, Kittler JT. J Biol Chem. 2014 May 23;289(21):14569-82. doi: 10.1074/jbc.M114.563031. Epub 2014 Mar 26. 10.1074/jbc.M114.563031 PubMed 24671417 |
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 pRK5-HA-Ubiquitin-K27R 4996 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Mammalian expression of HA tagged ubiquitin K27R.
ACCESSION .
VERSION .
KEYWORDS pRK5-HA-Ubiquitin-K27R
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4996)
AUTHORS Birsa N, Norkett R, Wauer T, Mevissen TE, Wu HC, Foltynie T, Bhatia
K, Hirst WD, Komander D, Plun-Favreau H, Kittler JT
TITLE Lysine 27 ubiquitination of the mitochondrial transport protein Miro
is dependent on serine 65 of the Parkin ubiquitin ligase.
JOURNAL J Biol Chem. 2014 May 23;289(21):14569-82. doi:
10.1074/jbc.M114.563031. Epub 2014 Mar 26.
PUBMED 24671417
REFERENCE 2 (bases 1 to 4996)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4996
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 1..17
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1..17
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
enhancer 46..425
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 426..629
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 580..600
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 626..650
/label=LNCX
/note="Human CMV promoter, forward primer"
promoter 860..878
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 860..877
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind 891..910
/label=pMT2-F
/note="Synthetic intron, forward primer"
regulatory 995..1004
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1007..1033
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 1064..1291
/codon_start=1
/gene="Homo sapiens UBC"
/product="human ubiquitin C"
/label=ubiquitin
/translation="MQIFVKTLTGKTITLEVEPSDTIENVRAKIQDKEGIPPDQQRLIF
AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
regulatory 1328..1337
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
polyA_signal 1341..1475
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1378..1397)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1432..1451
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
promoter 1544..1901
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(1567..1587)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 1752..1887
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 1814..1833
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(1921..1938)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(1921..1937)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1930..1952)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 2150..2605
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2237..2256)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2447..2468
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(2622..2641)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 2741..2763
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(2801..2819)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 2887..2991
/gene="bla"
/label=AmpR promoter
CDS 2992..3852
/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 complement(3210..3229)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 4023..4611
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4512..4531
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 4765..4782
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 4899..4920
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 4935..4965
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4973..4989
/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)."
primer_bind 4978..4
/label=M13/pUC Reverse
/note="In lacZ gene"
ORIGIN
1 caggaaacag ctatgacatg attacgaatt aattcgagct cgcccgacat tgattattga
61 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
121 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
181 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
241 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
301 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
361 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
421 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
481 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
541 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
601 tacggtggga ggtctatata agcagagctc gtttagtgaa ccgtcagatc gcctggagac
661 gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc ctccgcggcc
721 gggaacggtg cattggaacg cggattcccc gtgccaagag tgacgtaagt accgcctata
781 gagtctatag gcccaccccc ttggcttcgt tagaacgcgg ctacaattaa tacataacct
841 tatgtatcat acacatacga tttaggtgac actatagaat aacatccact ttgcctttct
901 ctccacaggt gtccactccc aggtccaact gcacctcggt tctatcgatt gaattcccga
961 accgacagtc ggtctcttca ccaaggccat tcgcgccacc atgggctacc cctatgatgt
1021 gcctgactac gcagatctca atggtggtgg tggtgggtcg accatgcaga tcttcgtgaa
1081 gaccctgact ggtaagacca tcactctcga agtggagccg agtgacacca ttgagaatgt
1141 cagggcaaag atccaagaca aggaaggcat ccctcctgac cagcagaggt tgatctttgc
1201 tgggaaacag ctggaagatg gacgcaccct gtctgactac aacatccaga aagagtccac
1261 cctgcacctg gtcctccgtc tcagaggtgg ttgagcggcc gctaagtaag taaggatccc
1321 cagcttggcc gccatggccc aacttgttta ttgcagctta taatggttac aaataaagca
1381 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
1441 ccaaactcat caatgtatct tatcatgtct ggatcgggaa ttaattcggc gcagcaccat
1501 ggcctgaaat aacctctgaa agaggaactt ggttaggtac cttctgaggc ggaaagaacc
1561 agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
1621 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
1681 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
1741 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
1801 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
1861 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctgtta acagcttggc
1921 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
1981 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
2041 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc ctgatgcggt attttctcct
2101 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt
2161 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
2221 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
2281 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
2341 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
2401 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
2461 caaactggaa caacactcaa ccctatctcg ggctattctt ttaatttata agggattttg
2521 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
2581 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc
2641 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
2701 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
2761 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
2821 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
2881 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
2941 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
3001 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
3061 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
3121 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
3181 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
3241 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
3301 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
3361 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
3421 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
3481 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
3541 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
3601 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
3661 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
3721 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
3781 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
3841 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
3901 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
3961 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
4021 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
4081 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
4141 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
4201 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
4261 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
4321 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
4381 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
4441 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
4501 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
4561 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
4621 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
4681 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
4741 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
4801 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
4861 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
4921 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
4981 ggataacaat ttcaca
//
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