| 质粒名称: | pcDNA GNSTM-3-RVG-10-Lamp2b-HA |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 71294 |
| 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中删除了一些限制性位点 |
| 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信号肽（N端插入）
GNSTM糖基化基序（N端插入）
狂犬病病毒糖蛋白（RVG）肽靶向乙酰胆碱受体（N末端插入）
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（克隆过程中销毁） |
| 3' 克隆位点: | BamHI（克隆过程中销毁） |
| 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 GNSTM-3-RVG-10-Lamp2b-HA was a gift from Joshua Leonard (Addgene plasmid # 71294 ; http://n2t.net/addgene:71294 ; RRID:Addgene_71294) |
| 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 GNSTM-3-RVG-10-Lamp2b-HA 6988 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Encodes (N to C): GNSTM glycosylation motif, 3 residue spacer, RVG
peptide, 10 residue spacer, Lamp2b (exosomal transmembrane protein),
3 residue spacer, HA tag..
ACCESSION .
VERSION .
KEYWORDS pcDNA GNSTM-3-RVG-10-Lamp2b-HA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6988)
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 6988)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6988
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 110..489
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 490..693
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 644..664
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 738..757
/label=T7
/note="T7 promoter, forward primer"
promoter 738..756
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
regulatory 779..788
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 2165..2191
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
primer_bind complement(2288..2305)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 2294..2518
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 2564..2992
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2651..2670)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2861..2882
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(3001..3021)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 3006..3335
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 3186..3321
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 3248..3267
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 3384..4409
/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 4539..4660
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(4576..4595)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 4630..4649
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(4709..4725)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4709..4725)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(4722..4744)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 4733..4749
/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(4757..4787)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4802..4823
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(4940..4957)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5111..5696)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5191..5210)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5867..6727)
/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 6490..6509
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6728..6832)
/gene="bla"
/label=AmpR promoter
primer_bind complement(6907..6926)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
61 gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg acattgatta
121 ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc atatatggag
181 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
241 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
301 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
361 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
421 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
481 attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
541 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
601 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
661 cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga acccactgct
721 tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg ctagtgccgc
781 cgccatggtg tgcttccgcc tcttcccggt tccgggctca gggctcgttc tggtctgcct
841 agtcctggga gctgtgcggt cttatgcagg taactcgact atgggcagtg gatacaccat
901 ttggatgccc gagaatccga gaccagggac accttgtgac atttttacca atagcagagg
961 gaagagagca tccaacgggg gcagtggatc tggatccggt ggctcgagtt tggaacttaa
1021 tttgacagat tcagaaaatg ccacttgcct ttatgcaaaa tggcagatga atttcacagt
1081 tcgctatgaa actacaaata aaacttataa aactgtaacc atttcagacc atggcactgt
1141 gacatataat ggaagcattt gtggggatga tcagaatggt cccaaaatag cagtgcagtt
1201 cggacctggc ttttcctgga ttgcgaattt taccaaggca gcatctactt attcaattga
1261 cagcgtctca ttttcctaca acactggtga taacacaaca tttcctgatg ctgaagataa
1321 aggaattctt actgttgatg aacttttggc catcagaatt ccattgaatg acctttttag
1381 atgcaatagt ttatcaactt tggaaaagaa tgatgttgtc caacactact gggatgttct
1441 tgtacaagct tttgtccaaa atggcacagt gagcacaaat gagttcctgt gtgataaaga
1501 caaaacttca acagtggcac ccaccataca caccactgtg ccatctccta ctacaacacc
1561 tactccaaag gaaaaaccag aagctggaac ctattcagtt aataatggca atgatacttg
1621 cctgctggct accatggggc tgcagctgaa catcactcag gataaggttg cttcagttat
1681 taacatcaac cccaatacaa ctcactccac aggcagctgc cgttctcaca ctgctctact
1741 tagactcaat agcagcacta ttaagtatct agactttgtc tttgctgtga aaaatgaaaa
1801 ccgattttat ctgaaggaag tgaacatcag catgtatttg gttaatggct ccgttttcag
1861 cattgcaaat aacaatctca gctactggga tgcccccctg ggaagttctt atatgtgcaa
1921 caaagagcag actgtttcag tgtctggagc atttcagata aatacctttg atctaagggt
1981 tcagcctttc aatgtgacac aaggaaagta ttctacagcc caagagtgtt cgctggatga
2041 tgacaccatt ctaatcccaa ttatagttgg tgctggtctt tcaggcttga ttatcgttat
2101 agtgattgct tacgtaattg gcagaagaaa aagttatgct ggatatcaga ctctgggaag
2161 cggataccca tacgatgtgc cagattacgc ttaaggatcc actagtccag tgtggtggaa
2221 ttctgcagat atccagcaca gtggcggccg ctcgagtcta gagggcccgt ttaaacccgc
2281 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
2341 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
2401 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
2461 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
2521 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
2581 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
2641 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
2701 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
2761 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
2821 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
2881 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
2941 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
3001 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
3061 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
3121 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
3181 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
3241 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
3301 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
3361 ttttcggatc tgatcagcac gtgatgaaaa agcctgaact caccgcgacg tctgtcgaga
3421 agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag
3481 aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct
3541 gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc
3601 cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc
3661 gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc
3721 agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt
3781 tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg
3841 cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt
3901 ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc
3961 acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag
4021 cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct
4081 tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc
4141 atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc
4201 aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat
4261 gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa
4321 gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc
4381 ccagcactcg tccgagggca aaggaatagc acgtgctacg agatttcgat tccaccgccg
4441 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
4501 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
4561 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
4621 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
4681 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
4741 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
4801 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
4861 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
4921 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
4981 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
5041 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
5101 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
5161 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
5221 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
5281 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
5341 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
5401 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
5461 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
5521 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
5581 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
5641 gtagcggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
5701 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
5761 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
5821 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
5881 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
5941 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
6001 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
6061 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
6121 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
6181 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
6241 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
6301 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
6361 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
6421 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
6481 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
6541 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
6601 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
6661 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
6721 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
6781 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
6841 cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac
6901 tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt
6961 gttggaggtc gctgagtagt gcgcgagc
//
