| 质粒名称: | pRK5-HA-Ubiquitin-K11 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 22901 |
| 存储实验室: | Sandra Weller |
|
载体骨架基本信息 |
|
| 载体名称: | pRK5-HA |
| 空载体大小: | 4800 bp |
| 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) |
| 插入大小: | 220 bp |
| 基因突变: | K11 only, all other lysines mutated to arginines |
| 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: | unknown (unknown if destroyed) |
| 3’克隆位点: | unknown (unknown if destroyed) |
| 5’测序引物 | SP6 |
|
文献引用方法 |
|
| 材料和方法部分: |
pRK5-HA-Ubiquitin-K11 was a gift from Sandra Weller (Addgene plasmid # 22901 ; http://n2t.net/addgene:22901 ; RRID:Addgene_22901)
|
| 参考文献部分: | Virus-Induced Chaperone-Enriched (VICE) domains function as nuclear protein quality control centers during HSV-1 infection. Livingston CM, Ifrim MF, Cowan AE, Weller SK. PLoS Pathog. 2009 Oct . 5(10):e1000619. 10.1371/journal.ppat.1000619 PubMed 19816571 |
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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-K11 4996 bp ds-DNA circular SYN 19-JAN-2022
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pRK5-HA-Ubiquitin-K11
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4996)
AUTHORS Livingston CM, Ifrim MF, Cowan AE, Weller SK
TITLE Virus-Induced Chaperone-Enriched (VICE) domains function as nuclear
protein quality control centers during HSV-1 infection.
JOURNAL PLoS Pathog. 2009 Oct . 5(10):e1000619.
PUBMED 19816571
REFERENCE 2 (bases 1 to 4996)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jan 19, 2022 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 complement(29..48)
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind 275..295
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
polyA_signal 387..521
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(411..430)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 465..484
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
regulatory 525..534
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS complement(571..798)
/codon_start=1
/product="human ubiquitin variant containing Lys-11, with
all other lysines mutated to arginine"
/label=ubiquitin-K11
/translation="MQIFVRTLTGKTITLEVEPSDTIENVRARIQDREGIPPDQQRLIF
AGRQLEDGRTLSDYNIQRESTLHLVLRLRGG"
CDS complement(829..855)
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
regulatory 858..867
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
primer_bind complement(952..971)
/label=pMT2-F
/note="Synthetic intron, forward primer"
promoter complement(984..1002)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(985..1002)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(1212..1236)
/label=LNCX
/note="Human CMV promoter, forward primer"
promoter complement(1233..1436)
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind complement(1262..1282)
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
enhancer 1437..1816
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
primer_bind complement(1845..1861)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1845..1861)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(1858..1880)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 1869..1885
/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(1893..1923)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 1938..1959
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(2076..2093)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2247..2835)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2327..2346)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(3006..3866)
/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 3629..3648
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(3867..3971)
/gene="bla"
/label=AmpR promoter
primer_bind 4039..4057
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(4095..4117)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 4217..4236
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
rep_origin complement(4253..4708)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4390..4411)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 4602..4621
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4906..4928
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 4920..4937
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 4921..4937
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter complement(4957..318)
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 4971..110
/label=SV40 ori
/note="SV40 origin of replication"
ORIGIN
1 ttctggaata gctcagaggc cgaggcggcc tcggcctctg cataaataaa aaaaattagt
61 cagccatggg gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt
121 aggggcggga ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct
181 ggggagcctg gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata
241 cttctgcctg ctggggagcc tggggacttt ccacacccta actgacacac attccacagc
301 tggttctttc cgcctcagaa ggtacctaac caagttcctc tttcagaggt tatttcaggc
361 catggtgctg cgccgaatta attcccgatc cagacatgat aagatacatt gatgagtttg
421 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta
481 ttgctttatt tgtaaccatt ataagctgca ataaacaagt tgggccatgg cggccaagct
541 ggggatcctt acttacttag cggccgctca accacctctt agtcttaaga caagatgtaa
601 ggtcgactct ctctgaatgt tgtaatcaga cagcgttcta ccgtcctcga gctgtctacc
661 ggcaaagatc aatctttgtt gatcaggtgg aatgccttct ctgtcttgaa ttctagctct
721 aacgttttcg atggtatcgg atggttcaac ttctagagtt atggttttac cggttaacgt
781 tctgacgaag atctgcatgg tcgacccacc accaccacca ttgagatctg cgtagtcagg
841 cacatcatag gggtagccca tggtggcgcg aatggccttg gtgaagagac cgactgtcgg
901 ttcgggaatt caatcgatag aaccgaggtg cagttggacc tgggagtgga cacctgtgga
961 gagaaaggca aagtggatgt tattctatag tgtcacctaa atcgtatgtg tatgatacat
1021 aaggttatgt attaattgta gccgcgttct aacgaagcca agggggtggg cctatagact
1081 ctataggcgg tacttacgtc actcttggca cggggaatcc gcgttccaat gcaccgttcc
1141 cggccgcgga ggctggatcg gtcccggtgt cttctatgga ggtcaaaaca gcgtggatgg
1201 cgtctccagg cgatctgacg gttcactaaa cgagctctgc ttatatagac ctcccaccgt
1261 acacgcctac cgcccatttg cgtcaatggg gcggagttgt tacgacattt tggaaagtcc
1321 cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa tggggtggag acttggaaat
1381 ccccgtgagt caaaccgcta tccacgccca ttgatgtact gccaaaaccg catcaccatg
1441 gtaatagcga tgactaatac gtagatgtac tgccaagtag gaaagtccca taaggtcatg
1501 tactgggcat aatgccaggc gggccattta ccgtcattga cgtcaatagg gggcgtactt
1561 ggcatatgat acacttgatg tactgccaag tgggcagttt accgtaaata ctccacccat
1621 tgacgtcaat ggaaagtccc tattggcgtt actatgggaa catacgtcat tattgacgtc
1681 aatgggcggg ggtcgttggg cggtcagcca ggcgggccat ttaccgtaag ttatgtaacg
1741 cggaactcca tatatgggct atgaactaat gaccccgtaa ttgattacta ttaataacta
1801 gtcaataatc aatgtcgggc gagctcgaat taattcgtaa tcatgtcata gctgtttcct
1861 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
1921 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
1981 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
2041 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
2101 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
2161 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
2221 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
2281 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
2341 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
2401 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
2461 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
2521 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
2581 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
2641 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
2701 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
2761 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
2821 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
2881 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
2941 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
3001 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
3061 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
3121 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
3181 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
3241 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
3301 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
3361 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
3421 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
3481 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
3541 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
3601 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
3661 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
3721 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
3781 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
3841 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
3901 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
3961 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc
4021 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt
4081 gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa
4141 gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg
4201 ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca taaaattgta
4261 aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
4321 caataggccg aaatcggcaa aatcccttat aaattaaaag aatagcccga gatagggttg
4381 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
4441 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc caaatcaagt
4501 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
4561 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
4621 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
4681 gcgcttaatg cgccgctaca gggcgcgtac tatggttgct ttgacgtatg cggtgtgaaa
4741 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc
4801 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
4861 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
4921 gtaaaacgac ggccagtgcc aagctgttaa cagctttttg caaaagccta ggcctccaaa
4981 aaagcctcct cactac
//
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