| 质粒名称: | pcDNA3.1(-) FLIPQTV3.0 100micro |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 63730 |
| 存储实验室: | Sakiko Okumoto |
|
载体骨架基本信息 |
|
| 载体名称: | pcDNA3.1 (-) |
| 载体出品公司: | invitrogen |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 5427 bp |
| 完整质粒大小: | 7528 bp |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | 新霉素（G418） |
| 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: | FLIPQTV3.0 sensor, 100microM affinity |
| 物种来源: | Synthetic |
| 插入大小: | 2119 bp |
| 基因突变: | D157N in glutamine binding domain |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pcDNA3.1(-) FLIPQTV3.0 100micro was a gift from Sakiko Okumoto (Addgene plasmid # 63730 ; http://n2t.net/addgene:63730 ; RRID:Addgene_63730) |
| 参考文献部分: | Visualization of glutamine transporter activities in living cells using genetically encoded glutamine sensors. Gruenwald K, Holland JT, Stromberg V, Ahmad A, Watcharakichkorn D, Okumoto S. PLoS One. 2012;7(6):e38591. doi: 10.1371/journal.pone.0038591. Epub 2012 Jun 14. 10.1371/journal.pone.0038591 PubMed 22723868 |
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pcDNA3.1(-) FLIPQTV3.0 100micro载体质粒基本信息
pcDNA3.1(-) FLIPQTV3.0 100micro质粒图谱载体图谱和pcDNA3.1(-) FLIPQTV3.0 100micro载体序列质粒序列多克隆位点信息
 FLIPQTV3.0 100micro质粒图谱.png)
pcDNA3.1(-) FLIPQTV3.0 100micro质粒载体简介
pcDNA3.1(-) FLIPQTV3.0 100micro质粒序列载体序列
LOCUS pcDNA3.1(-) FLIPQTV3.0 100micro 7528 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION FLIPQTV3.0 glutamine sensor with D157N mutation.
ACCESSION .
VERSION .
KEYWORDS pcDNA3.1(-) FLIPQTV3.0 100micro
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7528)
AUTHORS Gruenwald K, Holland JT, Stromberg V, Ahmad A, Watcharakichkorn D,
Okumoto S
TITLE Visualization of glutamine transporter activities in living cells
using genetically encoded glutamine sensors.
JOURNAL PLoS One. 2012;7(6):e38591. doi: 10.1371/journal.pone.0038591. Epub
2012 Jun 14.
PUBMED 22723868
REFERENCE 2 (bases 1 to 7528)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7528
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(44..63)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 235..614
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 615..818
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 769..789
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
primer_bind 863..882
/label=T7
/note="T7 promoter, forward primer"
promoter 863..881
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 1059..1766
/codon_start=1
/product="monomeric teal (cyan) fluorescent protein (Ai et
al., 2006)"
/label=mTFP1
/note="mammalian codon-optimized"
/translation="MVSKGEETTMGVIKPDMKIKLKMEGNVNGHAFVIEGEGEGKPYDG
TNTINLEVKEGAPLPFSYDILTTAFAYGNRAFTKYPDDIPNYFKQSFPEGYSWERTMTF
EDKGIVKVKSDISMEEDSFIYEIHLKGENFPPNGPVMQKKTTGWDASTERMYVRDGVLK
GDVKHKLLLEGGGHHRVDFKTIYRAKKAVKLPDYHFVDHRIEILNHDKDYNKVTVYESA
VARNSTDGMDELYK"
primer_bind complement(2421..2442)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind complement(2682..2701)
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
CDS 2841..3092
/codon_start=1
/product="C-terminal fragment of mVenus for use in
bimolecular fluorescence complementation (BiFC) (Kodama and
Hu, 2010)"
/label=VC155
/translation="DKQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNH
YLSYQSALSKDPNEKRDHMVLLEFVTAAGITLGMDELYK"
primer_bind 3029..3050
/label=EGFP-C
/note="EGFP, forward primer"
primer_bind complement(3122..3139)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 3128..3352
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 3398..3826
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3485..3504)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 3695..3716
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(3835..3855)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
promoter 3840..4169
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 4020..4155
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 4082..4101
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 4236..5030
/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
GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(4290..4309)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 4900..4919
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 5204..5325
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(5241..5260)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 5295..5314
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind complement(5374..5390)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5374..5390)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(5387..5409)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 5398..5414
/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(5422..5452)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 5467..5488
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5605..5622)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5776..6361)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(5856..5875)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(6532..7392)
/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 7155..7174
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(7393..7497)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
901 gtttaaacgg gccctctaga ctcgagcggc cgccactgtg ctggatatct gcagaattcc
961 accacactgg actagtggat ccactcgaga tggcaggctc gaaattagtt gtcgcgacgg
1021 ataccgcctt cgttccgttc gaatttaaac aggccccaat ggtgagcaag ggcgaggaga
1081 ccacaatggg cgtaatcaag cccgacatga agatcaagct gaagatggag ggcaacgtga
1141 atggccacgc cttcgtgatc gagggcgagg gcgagggcaa gccctacgac ggcaccaaca
1201 ccatcaacct ggaggtgaag gagggagccc ccctgccctt ctcctacgac attctgacca
1261 ccgcgttcgc ctacggcaac agggccttca ccaagtaccc cgacgacatc cccaactact
1321 tcaagcagtc cttccccgag ggctactctt gggagcgcac catgaccttc gaggacaagg
1381 gcatcgtgaa ggtgaagtcc gacatctcca tggaggagga ctccttcatc tacgagatac
1441 acctcaaggg cgagaacttc ccccccaacg gccccgtgat gcagaagaag accaccggct
1501 gggacgcctc caccgagagg atgtacgtgc gcgacggcgt gctgaagggc gacgttaagc
1561 acaagctgct gctggagggc ggcggccacc accgcgttga cttcaagacc atctacaggg
1621 ccaagaaggc ggtgaagctg cccgactatc actttgtgga ccaccgcatc gagatcctga
1681 accacgacaa ggactacaac aaggtgaccg tttacgagag cgccgtggcc cgcaactcca
1741 ccgacggcat ggacgagctg tacaaggtga gcgccaaata tgtgggcttt gacgtcgatc
1801 tgtgggctgc catcgctaaa gagctgaagc tggattacga actgaagccg atggatttca
1861 gtgggatcat tccggcactg caaaccaaaa acgtcgatct ggcgctggcg ggcattacca
1921 tcaccgacga gcgtaaaaaa gcgatcgatt tctctgacgg ctactacaaa agcggcctgt
1981 tagtgatggt gaaagctaac aataacgatg tgaaaagcgt gaaagatctc gacgggaaag
2041 tggttgctgt gaagagcggt actggctccg ttgattacgc gaaagcaaac atcaaaacta
2101 aagatctgcg tcagttcccg aacatcgata acgcctatat ggaactgggc accaaccgcg
2161 cagacgccgt tctgcacaac acgccaaaca ttctgtactt catcaaaacc gccggtaacg
2221 gtcagttcaa agcggtaggt gactctctgg aagcgcagca atacggtatt gcgttcccga
2281 aaggtagcga cgagctgcgt gacaaagtca acggcgcgtt gaaaaccctg cgcgagaacg
2341 gaacttacaa cgaaatctac aaaaaatggt tcggtactga accgaaaggc gctgagctgt
2401 tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca
2461 gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagctgatct
2521 gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg ggctacggcc
2581 tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca
2641 tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga
2701 cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca
2761 tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc
2821 acaacgtcta tatcaccgcc gacaagcaga agaacggcat caaggccaac ttcaagatcc
2881 gccacaacat cgaggacggc ggcgtgcagc tcgccgacca ctaccagcag aacaccccca
2941 tcggcgacgg ccccgtgctg ctgcccgaca atcactacct gagctaccag tccgccctga
3001 gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg
3061 ggatcactct cggcatggac gagctgtaca agtaaaagct taagtttaaa ccgctgatca
3121 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
3181 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
3241 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
3301 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag
3361 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta
3421 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
3481 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
3541 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
3601 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
3661 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
3721 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
3781 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg
3841 tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
3901 tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa
3961 gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta actccgccca
4021 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
4081 ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag
4141 gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg
4201 gatctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
4261 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
4321 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
4381 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
4441 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
4501 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
4561 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
4621 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
4681 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
4741 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
4801 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
4861 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
4921 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
4981 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
5041 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
5101 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
5161 cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc
5221 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
5281 actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc
5341 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
5401 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg
5461 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
5521 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
5581 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
5641 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
5701 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
5761 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
5821 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
5881 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
5941 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
6001 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
6061 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
6121 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
6181 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
6241 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
6301 cgctggtagc ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
6361 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
6421 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
6481 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
6541 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
6601 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
6661 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
6721 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
6781 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
6841 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
6901 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
6961 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
7021 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
7081 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
7141 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
7201 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
7261 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
7321 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
7381 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
7441 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
7501 atttccccga aaagtgccac ctgacgtc
//
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