| 质粒名称: | pcDNA Flag Lamp2b-HA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 71292 |
| 存储实验室: | Joshua Leonard |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA 3.1+ Hygro |
| 载体出品公司: | Clontech |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5578 bp |
| 完整质粒大小: | 6925 bp |
| 载体修饰: | 从MCS中删除了一些限制位点 |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 潮霉素 |
| 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: | Lamp2b |
| 物种来源: | H. sapiens (human) |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | CMV |
| 标签/融合蛋白: |
Lamp2 signal peptide（插入物上的N端）
Flag（插入N端子）
HA（插入件上的C端子）
|
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | NheI（在克隆过程中被破坏） |
| 3' 克隆位点: | BamHI（在克隆过程中被破坏） |
| 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: | pcDNA Flag Lamp2b-HA was a gift from Joshua Leonard (Addgene plasmid # 71292 ; http://n2t.net/addgene:71292 ; RRID:Addgene_71292) |
| 参考文献部分: | Stabilization of exosome-targeting peptides via engineered glycosylation. Hung ME, Leonard JN. J Biol Chem. 2015 Mar 27;290(13):8166-72. doi: 10.1074/jbc.M114.621383. Epub 2015 Feb 5. 10.1074/jbc.M114.621383 PubMed 25657008 |
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 pcDNA Flag Lamp2b-HA 6880 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Flag tag with 3 residue spacer fused to the N-terminus of Lamp2b, a
transmembrane exosomal protein. 3 residue spacer then HA tag fused
to the C-terminus of Lamp2b..
ACCESSION .
VERSION .
KEYWORDS pcDNA Flag Lamp2b-HA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6880)
AUTHORS Hung ME, Leonard JN
TITLE Stabilization of exosome-targeting peptides via engineered
glycosylation.
JOURNAL J Biol Chem. 2015 Mar 27;290(13):8166-72. doi:
10.1074/jbc.M114.621383. Epub 2015 Feb 5.
PUBMED 25657008
REFERENCE 2 (bases 1 to 6880)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6880
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin complement(160..745)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(240..259)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(916..1776)
/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 1539..1558
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(1777..1881)
/gene="bla"
/label=AmpR promoter
primer_bind complement(1956..1975)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 2147..2526
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 2527..2730
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 2681..2701
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 2775..2794
/label=T7
/note="T7 promoter, forward primer"
promoter 2775..2793
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 2816..2825
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2906..2929
/codon_start=1
/product="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/label=FLAG
/translation="DYKDDDDK"
CDS 4094..4120
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
primer_bind complement(4217..4234)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 4223..4447
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 4493..4921
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4580..4599)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4790..4811
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4930..4950)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 4935..5264
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 5115..5250
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 5177..5196
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 5313..6338
/codon_start=1
/gene="aph(4)-Ia"
/product="aminoglycoside phosphotransferase from E. coli"
/label=HygR
/note="confers resistance to hygromycin"
/translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
E"
polyA_signal 6468..6589
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(6505..6524)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 6559..6578
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(6638..6654)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(6638..6654)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(6651..6673)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 6662..6678
/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(6686..6716)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6731..6752
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(6869..6)
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
61 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
121 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
181 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
241 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
301 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
361 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
421 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
481 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
541 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
601 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
661 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtttt tttgtttgca agcagcagat
721 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
781 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
841 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
901 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
961 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
1021 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
1081 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
1141 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
1201 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
1261 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
1321 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
1381 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
1441 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
1501 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
1561 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
1621 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
1681 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
1741 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
1801 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
1861 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
1921 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
1981 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
2041 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
2101 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg
2161 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
2221 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
2281 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
2341 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
2401 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
2461 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
2521 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
2581 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
2641 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
2701 gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac
2761 tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta gtgccgccgc
2821 catggtgtgc ttccgcctct tcccggttcc gggctcaggg ctcgttctgg tctgcctagt
2881 cctgggagct gtgcggtctt atgcagacta taaagatgac gatgacaagg gctcgagttt
2941 ggaacttaat ttgacagatt cagaaaatgc cacttgcctt tatgcaaaat ggcagatgaa
3001 tttcacagtt cgctatgaaa ctacaaataa aacttataaa actgtaacca tttcagacca
3061 tggcactgtg acatataatg gaagcatttg tggggatgat cagaatggtc ccaaaatagc
3121 agtgcagttc ggacctggct tttcctggat tgcgaatttt accaaggcag catctactta
3181 ttcaattgac agcgtctcat tttcctacaa cactggtgat aacacaacat ttcctgatgc
3241 tgaagataaa ggaattctta ctgttgatga acttttggcc atcagaattc cattgaatga
3301 cctttttaga tgcaatagtt tatcaacttt ggaaaagaat gatgttgtcc aacactactg
3361 ggatgttctt gtacaagctt ttgtccaaaa tggcacagtg agcacaaatg agttcctgtg
3421 tgataaagac aaaacttcaa cagtggcacc caccatacac accactgtgc catctcctac
3481 tacaacacct actccaaagg aaaaaccaga agctggaacc tattcagtta ataatggcaa
3541 tgatacttgc ctgctggcta ccatggggct gcagctgaac atcactcagg ataaggttgc
3601 ttcagttatt aacatcaacc ccaatacaac tcactccaca ggcagctgcc gttctcacac
3661 tgctctactt agactcaata gcagcactat taagtatcta gactttgtct ttgctgtgaa
3721 aaatgaaaac cgattttatc tgaaggaagt gaacatcagc atgtatttgg ttaatggctc
3781 cgttttcagc attgcaaata acaatctcag ctactgggat gcccccctgg gaagttctta
3841 tatgtgcaac aaagagcaga ctgtttcagt gtctggagca tttcagataa atacctttga
3901 tctaagggtt cagcctttca atgtgacaca aggaaagtat tctacagccc aagagtgttc
3961 gctggatgat gacaccattc taatcccaat tatagttggt gctggtcttt caggcttgat
4021 tatcgttata gtgattgctt acgtaattgg cagaagaaaa agttatgctg gatatcagac
4081 tctgggaagc ggatacccat acgatgtgcc agattacgct taaggatcca ctagtccagt
4141 gtggtggaat tctgcagata tccagcacag tggcggccgc tcgagtctag agggcccgtt
4201 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
4261 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
4321 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
4381 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
4441 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
4501 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
4561 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
4621 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
4681 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
4741 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
4801 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
4861 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
4921 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
4981 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
5041 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
5101 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
5161 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
5221 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
5281 gtatatccat tttcggatct gatcagcacg tgatgaaaaa gcctgaactc accgcgacgt
5341 ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg
5401 agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg
5461 taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg
5521 ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt
5581 gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg
5641 ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga
5701 cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt
5761 tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg
5821 tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg
5881 aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc
5941 gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg
6001 ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg
6061 agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg
6121 gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc
6181 agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg
6241 cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa
6301 accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtgctacga gatttcgatt
6361 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
6421 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
6481 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
6541 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
6601 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
6661 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
6721 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
6781 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
6841 gtttgcgtat tgggcgctct tccgcttcct cgctcactga
//
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