| 质粒名称: | pRK5-HA-Ubiquitin-WT |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 17608 |
| 存储实验室: | Ted Dawson |
|
载体骨架基本信息 |
|
| 载体名称: | pRK5-HA |
| 载体抗性: | 氨苄霉素 |
| 空载体大小: | 4800 bp |
| 完整质粒大小: | 5020 bp |
| 载体类型: | 哺乳细胞表达 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Ubiquitin C |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 220 bp |
| 基因突变: | Ub |
| 标签/融合蛋白: | HA (N terminal on backbone) |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | SalI(未破坏) |
| 3' 克隆位点: | NotI (未破坏) |
| 5' 测序引物 | SP6 |
|
文献引用方法 |
|
| 材料和方法部分: | pRK5-HA-Ubiquitin-WT was a gift from Ted Dawson (爱迪基因 plasmid # 17608) |
| 参考文献部分: | Parkin mediates nonclassical, proteasomal-independent ubiquitination of synphilin-1: implications for Lewy body formation. Lim KL, Chew KC, Tan JM, Wang C, Chung KK, Zhang Y, Tanaka Y, Smith W, Engelender S, Ross CA, Dawson VL, Dawson TM. J Neurosci. 2005 Feb 23. 25(8):2002-9. 10.1523/JNEUROSCI.4474-04.2005 PubMed 15728840 |
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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-WT 4996 bp ds-DNA circular SYN 10-JAN-2018
DEFINITION Mammalian expression of HA tagged ubiquitin.
ACCESSION .
VERSION .
KEYWORDS pRK5-HA-Ubiquitin-WT
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4996)
AUTHORS Lim KL, Chew KC, Tan JM, Wang C, Chung KK, Zhang Y, Tanaka Y, Smith
W, Engelender S, Ross CA, Dawson VL, Dawson TM
TITLE Parkin mediates nonclassical, proteasomal-independent ubiquitination
of synphilin-1: implications for Lewy body formation.
JOURNAL J Neurosci. 2005 Feb 23. 25(8):2002-9.
PUBMED 15728840
COMMENT This file is created by Vector NTI
http://www.biofeng.com/
COMMENT ORIGDB|GenBank
COMMENT VNTAUTHORNAME|biofeng.com|
COMMENT VNTNAME|pRK5-HA-Ubiquitin-WT|
FEATURES Location/Qualifiers
source 1..4996
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 1..380
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 381..584
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 535..555
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 581..605
/label=LNCX
/note="Human CMV promoter, forward primer"
promoter 815..833
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 815..832
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind 846..865
/label=pMT2-F
/note="Synthetic intron, forward primer"
regulatory 950..959
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 962..988
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 1019..1246
/codon_start=1
/gene="Homo sapiens UBC"
/product="human ubiquitin C"
/label=ubiquitin
/translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
regulatory 1283..1292
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
polyA_signal 1296..1430
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1333..1352)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1387..1406
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
promoter 1499..1856
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(1522..1542)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 1707..1842
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 1769..1788
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(1876..1893)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(1876..1892)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1885..1907)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 2105..2560
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2192..2211)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2402..2423
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(2577..2596)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 2696..2718
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(2756..2774)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 2842..2946
/gene="bla"
/label=AmpR promoter
CDS 2947..3807
/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(3165..3184)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 3978..4566
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4467..4486
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 4720..4737
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 4854..4875
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 4890..4920
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4928..4944
/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 4933..4955
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 4952..4968
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 4952..4968
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
ORIGIN
1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
601 agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagacac cgggaccgat
661 ccagcctccg cggccgggaa cggtgcattg gaacgcggat tccccgtgcc aagagtgacg
721 taagtaccgc ctatagagtc tataggccca cccccttggc ttcgttagaa cgcggctaca
781 attaatacat aaccttatgt atcatacaca tacgatttag gtgacactat agaataacat
841 ccactttgcc tttctctcca caggtgtcca ctcccaggtc caactgcacc tcggttctat
901 cgattgaatt cccgaaccga cagtcggtct cttcaccaag gccattcgcg ccaccatggg
961 ctacccctat gatgtgcctg actacgcaga tctcaatggt ggtggtggtg ggtcgaccat
1021 gcagatcttc gtgaagaccc tgactggtaa gaccatcact ctcgaagtgg agccgagtga
1081 caccattgag aatgtcaagg caaagatcca agacaaggaa ggcatccctc ctgaccagca
1141 gaggttgatc tttgctggga aacagctgga agatggacgc accctgtctg actacaacat
1201 ccagaaagag tccaccctgc acctggtcct ccgtctcaga ggtggttgag cggccgctaa
1261 gtaagtaagg atccccagct tggccgccat ggcccaactt gtttattgca gcttataatg
1321 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
1381 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc gggaattaat
1441 tcggcgcagc accatggcct gaaataacct ctgaaagagg aacttggtta ggtaccttct
1501 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
1561 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
1621 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
1681 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
1741 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
1801 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
1861 tgttaacagc ttggcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt
1921 tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga
1981 ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat ggcgcctgat
2041 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
2101 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
2161 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
2221 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
2281 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
2341 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
2401 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcgggcta ttcttttaat
2461 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
2521 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
2581 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
2641 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
2701 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
2761 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
2821 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
2881 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
2941 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
3001 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
3061 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
3121 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
3181 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
3241 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
3301 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
3361 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
3421 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
3481 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
3541 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
3601 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
3661 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
3721 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
3781 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
3841 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
3901 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
3961 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
4021 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
4081 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
4141 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
4201 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
4261 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
4321 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
4381 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
4441 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
4501 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
4561 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
4621 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
4681 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
4741 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
4801 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
4861 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
4921 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacat gattacgaat
4981 taattcgagc tcgccc
//
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