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- 详细信息
- 文献和实验
- 技术资料
- 保存条件:
-20
- 保质期:
2年
- 英文名:
ADRA2; ADRAR; ZNF32; ADRA2R; ALPHA2A
- 库存:
100
- 供应商:
上海烜雅生物科技有限公司
- 规格:
干粉/液体
名称:pEGFP-ADRA2A人源基因质粒
别称: ADRA2; ADRAR; ZNF32; ADRA2R; ALPHA2A
启动子:CMV
复制子: pUC
终止子: SV40 poly(A) signal
质粒大小:6095bp
质粒标签:C-EGFP
原核抗性: Kan
筛选标记:G418
克隆菌株: DH5a
培养条件: 37度
质粒属性
载体宿主:哺乳细胞
载体用途:蛋白表达
基因种属:人
基因类型:ORF
原核抗性:Kan
真核抗性:Neo/G418
荧光蛋白:
质粒简介
pEGFP-ADRA2A质粒是一个人源基因过表达质粒,CMV启动子驱动绿色荧光蛋白EGFP和人源基因ADRA2A融合在一起表达。
质粒图谱
质粒序列
LOCUS Exported 6095 bp ds-DNA circular SYN 02-SEP-2017
DEFINITION synthetic circular DNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6095)
AUTHORS admin
TITLE Direct Submission
http://www.miaolingbio.com
REFERENCE 2 (bases 1 to 6095)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Oct 24, 2017 from SnapGene Viewer 4.0.7
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6095
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 61..364
/label=CMV enhancer
/note="CMV enhancer;human cytomegalovirus immediate early
enhancer"
promoter 365..568
/label=CMV promoter
/note="CMV promoter;human cytomegalovirus (CMV) immediate
early promoter"
misc_feature 625..2019
/label=ADRA2A
/note="Homo sapiens adrenoceptor alpha 2A (ADRA2A), mRNA
NCBI Reference Sequence: NM_000681.3"
CDS 2041..2760
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/note="EGFP;mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
polyA_signal 2883..3004
/label=SV40 poly(A) signal
/note="SV40 poly(A) signal;SV40 polyadenylation signal"
rep_origin complement(3011..3466)
/direction=LEFT
/label=f1 ori
/note="f1 ori;f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 3493..3597
/gene="bla"
/label=bla AmpR promoter
/note="AmpR promoter"
promoter 3599..3956
/label=SV40 promoter
/note="SV40 promoter;SV40 enhancer and early promoter"
rep_origin 3807..3942
/label=SV40 ori
/note="SV40 ori;SV40 origin of replication"
CDS 3991..4785
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="NeoR/KanR;confers resistance to neomycin, kanamycin,
and G418 (Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPAT-CPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 5017..5064
/label=HSV TK poly(A) signal
/note="HSV TK poly(A) signal;herpesvirus thymidine kinase
polyadenylation signal"
rep_origin 5393..5981
/direction=RIGHT
/label=ori
/note="ori;high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
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 atctcgaggc caccatgttc cgccaggagc agccgttggc cgagggcagc
661 tttgcgccca tgggctccct gcagccggac gcgggcaacg cgagctggaa cgggaccgag
721 gcgccggggg gcggcgcccg ggccacccct tactccctgc aggtgacgct gacgctggtg
781 tgcctggccg gcctgctcat gctgctcacc gtgttcggca acgtgctcgt catcatcgcc
841 gtgttcacga gccgcgcgct caaggcgccc caaaacctct tcctggtgtc tctggcctcg
901 gccgacatcc tggtggccac gctcgtcatc cctttctcgc tggccaacga ggtcatgggc
961 tactggtact tcggcaaggc ttggtgcgag atctacctgg cgctcgacgt gctcttctgc
1021 acgtcgtcca tcgtgcacct gtgcgccatc agcctggacc gctactggtc catcacacag
1081 gccatcgagt acaacctgaa gcgcacgccg cgccgcatca aggccatcat catcaccgtg
1141 tgggtcatct cggccgtcat ctccttcccg ccgctcatct ccatcgagaa gaagggcggc
1201 ggcggcggcc cgcagccggc cgagccgcgc tgcgagatca acgaccagaa gtggtacgtc
1261 atctcgtcgt gcatcggctc cttcttcgct ccctgcctca tcatgatcct ggtctacgtg
1321 cgcatctacc agatcgccaa gcgtcgcacc cgcgtgccac ccagccgccg gggtccggac
1381 gccgtcgccg cgccgccggg gggcaccgag cgcaggccca acggtctggg ccccgagcgc
1441 agcgcgggcc cggggggcgc agaggccgaa ccgctgccca cccagctcaa cggcgcccct
1501 ggcgagcccg cgccggccgg gccgcgcgac accgacgcgc tggacctgga ggagagctcg
1561 tcttccgacc acgccgagcg gcctccaggg ccccgcagac ccgagcgcgg tccccggggc
1621 aaaggcaagg cccgagcgag ccaggtgaag ccgggcgaca gcctgccgcg gcgcgggccg
1681 ggggcgacgg ggatcgggac gccggctgca gggccggggg aggagcgcgt cggggctgcc
1741 aaggcgtcgc gctggcgcgg gcggcagaac cgcgagaagc gcttcacgtt cgtgctggcc
1801 gtggtcatcg gagtgttcgt ggtgtgctgg ttccccttct tcttcaccta cacgctcacg
1861 gccgtcgggt gctccgtgcc acgcacgctc ttcaaattct tcttctggtt cggctactgc
1921 aacagctcgt tgaacccggt catctacacc atcttcaacc acgatttccg ccgcgccttc
1981 aagaagatcc tctgtcgggg ggacaggaag cggatcgtga cggatccacc ggtcgccacc
2041 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
2101 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
2161 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
2221 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
2281 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
2341 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
2401 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
2461 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
2521 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
2581 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
2641 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
2701 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa
2761 agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt ttacttgctt
2821 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
2881 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
2941 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
3001 cttaaggcgt aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
3061 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
3121 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
3181 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
3241 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
3301 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
3361 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
3421 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt
3481 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
3541 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtcct
3601 gaggcggaaa gaaccagctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
3661 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
3721 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
3781 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
3841 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctcggcct
3901 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaagatc
3961 gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt gcacgcaggt
4021 tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca gacaatcggc
4081 tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct ttttgtcaag
4141 accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct atcgtggctg
4201 gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc gggaagggac
4261 tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct tgctcctgcc
4321 gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga tccggctacc
4381 tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg gatggaagcc
4441 ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc agccgaactg
4501 ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat
4561 gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat cgactgtggc
4621 cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga tattgctgaa
4681 gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat
4741 tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg actctggggt
4801 tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
4861 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
4921 agcgcgggga tctcatgctg gagttcttcg cccaccctag ggggaggcta actgaaacac
4981 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
5041 cgcacggtgt tgggtcgttt gttcataaac gcggggttcg gtcccagggc tggcactctg
5101 tcgatacccc accgagaccc cattggggcc aatacgcccg cgtttcttcc ttttccccac
5161 cccacccccc aagttcgggt gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc
5221 ctgccatagc ctcaggttac tcatatatac tttagattga tttaaaactt catttttaat
5281 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
5341 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
5401 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
5461 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
5521 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
5581 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
5641 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
5701 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
5761 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
5821 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
5881 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
5941 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
6001 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
6061 ctgattctgt ggataaccgt attaccgcca tgcat
//
质粒菌株产品操作说明书
一、扩增流程
收到产品后,请先根据产品管壁标签来判断产品形式,并在扩增前准确查找该质粒菌株的抗性、感受态和培养温度。
1、质粒干粉(常温运输,存于-20度,90天保质期,请务必转化挑单克隆培养,不要直接使用和测序)
①收到质粒干粉后请先5000rpm离心1min,再加入20μl ddH2O去离子水溶解质粒;
②取1支100μl 感受态于冰上解冻10min,加入2μl质粒,再冰浴30min后,42℃热激60s,不要搅动,再冰浴2min;(从第二步开始均要在超净工作台中无菌操作)
③加入900μl无抗的LB液体培养基,180rpm震荡37℃培养45min;
④6000rpm离心5min,仅留100μl上清液重悬细菌沉淀,并涂布至目标质粒抗性的LB平板上;(可使用本平台的平板涂布专用玻璃珠进行涂布,可以比传统涂布方法获得更多转化子)
⑤将平板正向培养1h,再倒置37℃培养14h。如果要求是30度则培养20h;
(菌落过多则将质粒稀释后再转化。没有菌落则加入10μl质粒转化。另不要直接转表达感受态,要先转克隆感受态,重提质粒后再导入表达感受态)
⑥挑取单菌落至LB液体培养基中,加入对应抗生素,220rpm震荡培养14h,根据实验需要和质粒提取试剂盒说明书提取质粒。
2、甘油菌种(冰袋运输,存于-80℃,保质期90天,请务必划线挑单克隆培养)
四区划线后挑单菌落培养,酵母菌需要先液体复苏再四区划线,再挑单菌落液体培养。
3、穿刺菌种(冰袋运输,存于4℃,保质期7天)
穿刺接种,液体培养后四区划线,再挑单菌落液体培养。
4、菌落平板(冰袋运输,存于4℃,保质期7天)
直接挑取单菌落至液体培养基中。
5、液体质粒(冰袋运输,存于-20℃,保质期90天)
单独提取的液体质粒收到后可直接使用。
6、滤纸质粒(常温运输,存于-20度,90天保质期,请务必转化挑单克隆培养,不要直接使用和测序)
收到货后将滤纸画圈部分剪下放入EP管中,加100ul无菌水将滤纸浸湿并浸泡5min,吸取5ul质粒转化,离心全涂。
二、转化图片
| P4040/pCMV-SPORT6-ARMCX2(1同义突变)人源基因质粒 |
| P4041/pCMV-SPORT6-TMEM183A人源基因质粒 |
| P4042/pCMV-SPORT6-LRIF1(603-2310bp)人源基因质粒 |
| P4043/pCMV-SPORT6-GRAMD2B人源基因质粒 |
| P4044/pCMV-SPORT6-OXCT1人源基因质粒 |
| P4045/pCMV-SPORT6-UTP15(1同义突变)人源基因质粒 |
| P4046/pCMV-SPORT6-RPS3A人源基因质粒 |
| P4047/pCMV-SPORT6-RAB38(1同义突变)人源基因质粒 |
| P4048/pCMV-SPORT6-ABRAXAS2人源基因质粒 |
| P4049/pCMV-SPORT6-SMNDC1人源基因质粒 |
| P4050/pCMV-SPORT6-C11orf68人源基因质粒 |
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文献和实验Restriction Map and Multiple Cloning Site of pEGFP-N2. (Unique restriction sites are in color or bold.) The Not I site follows the EGFP stop codon. The Nhe I site cannot be used for fusions since it contains an in-frame stop codon
多克隆位点区 Restriction Map and Multiple Cloning Site of pEGFP-N1. (Unique restriction sites are in color or bold.) The Not I site follows the EGFP stop codon. The Xba I site (*) is methylated in the DNA provided by CLONTECH. If you wish to digest
多克隆位点区(MCS) Restriction Map and Multiple Cloning Site of pEGFP-C2. (Unique restriction sites are in color or bold.) Note that the Eag I site is not unique.) The Xba I site cannot be used for fusions











