| 質粒名稱: | PH-Akt-GFP |
|---|---|
| 出品公司: | Addgene |
| 目錄編號: | 51465 |
| 存儲實驗室: | Tamas Balla |
|
載體骨架基本信息 |
|
| 載體名稱: | pEGFP-N1 |
| 載體出品公司: | Clontech |
| 空載體大小: | 5286bp |
| 完整質粒大小: | 4989 bp |
| 載體類型: | 哺乳細胞表達 |
| 篩選標記: | 新霉素（G418） |
| 載體抗性: | Kanamycin, 50 μg/mL |
| 生長溫度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷貝數: | 高拷貝 |
|
插入基因信息 |
|
| 插入基因名稱: | AKT (aa 1–164) |
| 插入大小: | 492 bp |
| GenBank編號 | NM_001014431.1 |
| 啟動子: | CMV |
| 標簽/融合蛋白: | EGFP (C terminal on insert) |
| 克隆方法: | 限制性內切酶 |
| 5’克隆位點: | EcoRI (not destroyed) |
| 3’克隆位點: | BamHI (not destroyed) |
| 5’測序引物 | CMV-F |
| 3’測序引物 | EGFP-N |
|
文獻引用方法 |
|
| 材料和方法部分: | PH-Akt-GFP was a gift from Tamas Balla (Addgene plasmid # 51465 ; //n2t.net/addgene:51465 ; RRID:Addgene_51465) |
| 參考文獻部分: | Visualization of phosphoinositides that bind pleckstrin homology domains: calcium- and agonist-induced dynamic changes and relationship to myo-[3H]inositol-labeled phosphoinositide pools. Varnai P, Balla T. J Cell Biol. 1998 Oct 19;143(2):501-10. 10.1083/jcb.143.2.501 PubMed 9786958 |
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PH-Akt-GFP載體質粒基本信息
PH-Akt-GFP質粒圖譜載體圖譜和PH-Akt-GFP載體序列質粒序列多克隆位點信息
PH-Akt-GFP質粒載體簡介
PH-Akt-GFP質粒序列載體序列
LOCUS PH-Akt-GFP 5286 bp DNA circular SYN 14-APR-2025
DEFINITION Is a biosensor for PIP3/PI(3,4)P2.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5286)
AUTHORS Varnai P, Balla T
TITLE Visualization of phosphoinositides that bind pleckstrin homology
domains: calcium- and agonist-induced dynamic changes and
relationship to myo-[3H]inositol-labeled phosphoinositide pools.
JOURNAL J Cell Biol. 1998 Oct 19;143(2):501-10.
PUBMED 9786958
REFERENCE 2 (bases 1 to 5286)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Apr 14, 2025 from SnapGene Server 8.0.1
//gt12365.cn
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J Cell
Biol."; date: "1998-10-19"; volume: "143(2)"; pages: "501-10"
COMMENT Sequence Label: PH-Akt-GFP
FEATURES Location/Qualifiers
source 1..5286
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 61..364
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 365..568
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 519..539
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
regulatory 1226..1235
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
CDS join(1232..1234,1235..1237,1238..1951)
/codon_start=1
/product="the original enhanced GFP (Yang et al., 1996)"
/label=EGFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS join(1232..1234,1235..1237,1238..1948)
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(1277..1298)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(1538..1557)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 1885..1906
/label=EGFP-C
/note="EGFP, forward primer"
polyA_signal 2074..2195
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2111..2130)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 2165..2184
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin complement(2202..2657)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2339..2360)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 2551..2570
/label=F1ori-R
/note="F1 origin, reverse primer"
promoter 2684..2788
/gene="bla"
/label=AmpR promoter
promoter 2790..3147
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
enhancer 2790..2981
/label=SV40 enhancer
/note="enhancer for the SV40 early promoter (Herr, 1993)"
primer_bind complement(2813..2833)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 2998..3133
/label=SV40 ori
/note="SV40 origin of replication"
CDS 3182..3976
/codon_start=1
/product="aminoglycoside phosphotransferase"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
CDS 3182..3976
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(3236..3255)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 3846..3865
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
primer_bind complement(4164..4183)
/label=TK-pA-R
/note="Thymidine kinase polyA, reverse primer"
polyA_signal 4208..4255
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 4584..5172
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 5073..5092
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
601 ccggactcag atctcgagct caagcttcga attctgcatc agaggctgtg gccaggccag
661 ctgggctcgg ggagcgccag cctgagagga gcgcgtgagc gtcgcgggag cctcgggcac
721 catgagcgac gtggctattg tgaaggaggg ttggctgcac aaacgagggg agtacatcaa
781 gacctggcgg ccacgctact tcctcctcaa gaatgatggc accttcattg gctacaagga
841 gcggccgcag gatgtggacc aacgtgaggc tcccctcaac aacttctctg tggcgcagtg
901 ccagctgatg aagacggagc ggccccggcc caacaccttc atcatccgct gcctgcagtg
961 gaccactgtc atcgaacgca ccttccatgt ggagactcct gaggagcggg aggagtggac
1021 aaccgccatc cagactgtgg ctgacggcct caagaagcag gaggaggagg agatggactt
1081 ccggtcgggc tcacccagtg acaactcagg ggctgaagag atggaggtgt ccctggccaa
1141 gcccaagcac cgcgtgacca tgaacgagtt tgagtacctg aagctgctgg gcaagggcac
1201 tttcggcaag gtggatccac cggtcgccac catggtgagc aagggcgagg agctgttcac
1261 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
1321 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
1381 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca
1441 gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
1501 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
1561 cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
1621 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
1681 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
1741 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
1801 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa
1861 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
1921 cactctcggc atggacgagc tgtacaagta aagcggccgc gactctagat cataatcagc
1981 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
2041 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
2101 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
2161 agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa gcgttaatat
2221 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
2281 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
2341 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
2401 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
2461 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
2521 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
2581 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
2641 gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
2701 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
2761 gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg
2821 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
2881 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
2941 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
3001 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
3061 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
3121 ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg
3181 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
3241 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
3301 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
3361 gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
3421 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
3481 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
3541 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
3601 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
3661 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga
3721 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
3781 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
3841 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
3901 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
3961 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
4021 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
4081 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
4141 gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
4201 gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt tgttcataaa
4261 cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc
4321 caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca
4381 gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta ctcatatata
4441 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
4501 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
4561 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
4621 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
4681 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
4741 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
4801 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
4861 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
4921 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
4981 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
5041 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
5101 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
5161 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
5221 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
5281 atgcat
//
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