pmirGLO哺乳报告质粒价格_品牌:Xybio-丁香通官网

保存条件：-20

保质期：2年

英文名：pmir-GLO

库存：100

供应商：上海烜雅生物科技有限公司

CAS号：/

规格：干粉/液体

基本信息
名称：pmirGLO哺乳报告质粒
别称: pmir-GLO

启动子:	PGK
复制子:	pUC
终止子:	SV40 poly(A) signal
质粒分类:	哺乳细胞，信号通路报告载体
质粒大小:	7350bp
原核抗性:	Amp
筛选标记:	G418
克隆菌株:	DH5a
培养条件:	37度
表达宿主:	哺乳细胞
诱导方式:	无须诱导，瞬时表达
5'测序引物:	PmirGLO-F：GACGAGGTGCCTAAAGGACT
3'测序引物:	根据序列设计引物

质粒属性

载体宿主：	哺乳细胞
载体用途：	信号报告
基因种属：	空载体
基因类型：	miRNA,UTR
原核抗性：	Amp
真核抗性：	Neo/G418
荧光蛋白：	fLuc,rLuc

质粒简介
The pmirGLO Dual-Luciferase miRNA Target Expression Vector(a–d) is designed to quantitatively evaluate microRNA (miRNA) activity by the insertion of miRNA target sites 3´ of the firefly luciferase gene (luc2). These target sites can be introduced by cloning putative miRNA binding sites alone, or the 3´ untranslated region (UTR) of a gene of interest, to study the influence of these sites on transcript stability and activity. Firefly luciferase is the primary reporter gene; reduced firefly luciferase expression indicates the binding of endogenous or introduced miRNAs to the cloned miRNA target sequence. This vector is based on Promega dual-luciferase technology, with firefly luciferase (luc2) used as the primary reporter to monitor mRNA regulation and Renilla luciferase (hRluc-neo) acting as a control reporter for normalization and selection.

质粒图谱

质粒序列
LOCUS Exported 7350 bp ds-DNA circular SYN 04-SEP-2016
DEFINITION synthetic circular DNA
KEYWORDS Untitled
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7350)
AUTHORS .
TITLE Direct Submission
http://www.miaolingbio.com
FEATURES Location/Qualifiers
source 1..7350
/organism="synthetic DNA construct"
/mol_type="other DNA"
polyA_signal 115..236
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
promoter 481..838
/note="SV40 promoter"
/note="SV40 enhancer and early promoter"
rep_origin 689..824
/note="SV40 ori"
/note="SV40 origin of replication"
CDS 889..1821
/codon_start=1
/product="Renilla luciferase"
/note="hRluc"
/note="human codon-optimized"
/translation="MASKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
CDS 1873..2664
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/note="NeoR/KanR"
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
HLAPAEKVSIMADAMRRLHTLDPATC-PFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 2728..2776
/note="synthetic polyadenylation signal"
promoter 2932..3036
/gene="bla"
/note="AmpR promoter"
CDS 3037..3897
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/note="AmpR"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin complement(4055..4643)
/direction=LEFT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature 4759..5042
/note="cer region"
/note="ColE1-derived recombination site that helps to
maintain plasmids as monomers"
promoter 5099..5598
/note="PGK promoter"
/note="mouse phosphoglycerate kinase 1 promoter"
CDS 5645..7297
/codon_start=1
/product="firefly luciferase"
/note="luciferase"
/note="synthetic luc2 version of the luciferase gene"
/translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
ORIGIN
1 catgcaagct gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
61 cgctgagcaa taactagcat aaccccttgg ggcggccgct tcgagcagac atgataagat
121 acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg
181 aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca
241 acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag gtttttttaa
301 gcaagtaaaa cctctacaaa tgtggtaaaa tcgaatttta acaaaatatt aacgcttaca
361 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgcg
421 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt taggtacctt
481 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
541 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
601 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
661 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
721 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
781 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
841 gcttgattct tctgacacaa cagtctcgaa ccaaaggctg gagccaccat ggcttccaag
901 gtgtacgacc ccgagcaacg caaacgcatg atcactgggc ctcagtggtg ggctcgctgc
961 aagcaaatga acgtgctgga ctccttcatc aactactatg attccgagaa gcacgccgag
1021 aacgccgtga tttttctgca tggtaacgct gcctccagct acctgtggag gcacgtcgtg
1081 cctcacatcg agcccgtggc tagatgcatc atccctgatc tgatcggaat gggtaagtcc
1141 ggcaagagcg ggaatggctc atatcgcctc ctggatcact acaagtacct caccgcttgg
1201 ttcgagctgc tgaaccttcc aaagaaaatc atctttgtgg gccacgactg gggggcttgt
1261 ctggcctttc actactccta cgagcaccaa gacaagatca aggccatcgt ccatgctgag
1321 agtgtcgtgg acgtgatcga gtcctgggac gagtggcctg acatcgagga ggatatcgcc
1381 ctgatcaaga gcgaagaggg cgagaaaatg gtgcttgaga ataacttctt cgtcgagacc
1441 atgctcccaa gcaagatcat gcggaaactg gagcctgagg agttcgctgc ctacctggag
1501 ccattcaagg agaagggcga ggttagacgg cctaccctct cctggcctcg cgagatccct
1561 ctcgttaagg gaggcaagcc cgacgtcgtc cagattgtcc gcaactacaa cgcctacctt
1621 cgggccagcg acgatctgcc taagatgttc atcgagtccg accctgggtt cttttccaac
1681 gctattgtcg agggagctaa gaagttccct aacaccgagt tcgtgaaggt gaagggcctc
1741 cacttcagcc aggaggacgc tccagatgaa atgggtaagt acatcaagag cttcgtggag
1801 cgcgtgctga agaacgagca gaccggtggt gggagcggag gtggcggatc aggtggcgga
1861 ggctccggag ggattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
1921 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
1981 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
2041 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
2101 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
2161 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
2221 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
2281 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
2341 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
2401 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
2461 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
2521 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
2581 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
2641 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
2701 cgcccaacct gccatcacga tggccgcaat aaaatatctt tattttcatt acatctgtgt
2761 gttggttttt tgtgtgaatc gatagcgata aggatcctct ttgcgcttgc gttttccctt
2821 gtccagatag cccagtagct gacattcatc cggggtcagc accgtttctg cggactggct
2881 ttctacgtaa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
2941 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
3001 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
3061 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
3121 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
3181 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
3241 actttcaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
3301 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
3361 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
3421 gataacactg cggccaactt acttctgaca actatcggag gaccgaagga gctaaccgct
3481 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
3541 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
3601 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
3661 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
3721 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
3781 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
3841 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaattc
3901 gaaatgaccg accaagcgac gcccaaccgg tatcagctca ctcaaaggcg gtaatacggt
3961 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
4021 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
4081 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
4141 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
4201 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
4261 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
4321 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
4381 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
4441 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
4501 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
4561 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
4621 cgcgcagaaa aaaaggattt caagaagatc ctttgatctt ttctacgggg tctgacgctc
4681 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
4741 cctagatcct tttatagtcc ggaaatacag gaacgcacgc tggatggccc ttcgctggga
4801 tggtgaaacc atgaaaaatg gcagcttcag tggattaagt gggggtaatg tggcctgtac
4861 cctctggttg cataggtatt catacggtta aaatttatca ggcgcgattg cggcagtttt
4921 tcgggtggtt tgttgccatt tttacctgtc tgctgccgtg atcgcgctga acgcgtttta
4981 gcggtgcgta caattaaggg attatggtaa atccacttac tgtctgccct cgtagccatc
5041 gagataaacc gcagtactcc ggccacgatg cgtccggcgt agaggatcga gatctaccgg
5101 gtaggggagg cgcttttccc aaggcagtct ggagcatgcg ctttagcagc cccgctgggc
5161 acttggcgct acacaagtgg cctctggcct cgcacacatt ccacatccac cggtaggcgc
5221 caaccggctc cgttctttgg tggccccttc gcgccacctt ctactcctcc cctagtcagg
5281 aagttccccc ccgccccgca gctcgcgtcg tgcaggacgt gacaaatgga agtagcacgt
5341 ctcactagtc tcgtgcagat ggacagcacc gctgagcaat ggaagcgggt aggcctttgg
5401 ggcagcggcc aatagcagct ttgctccttc gctttctggg ctcagaggct gggaaggggt
5461 gggtccgggg gcgggctcag gggcgggctc aggggcgggg cgggcgcccg aaggtcctcc
5521 ggaggcccgg cattctgcac gcttcaaaag cgcacgtctg ccgcgctgtt ctcctcttcc
5581 tcatctccgg gcctttcgac ctgcagccca agcttggcaa tccggtactg ttggtaaagc
5641 caccatggaa gatgccaaaa acattaagaa gggcccagcg ccattctacc cactcgaaga
5701 cgggaccgcc ggcgagcagc tgcacaaagc catgaagcgc tacgccctgg tgcccggcac
5761 catcgccttt accgacgcac atatcgaggt ggacattacc tacgccgagt acttcgagat
5821 gagcgttcgg ctggcagaag ctatgaagcg ctatgggctg aatacaaacc atcggatcgt
5881 ggtgtgcagc gagaatagct tgcagttctt catgcccgtg ttgggtgccc tgttcatcgg
5941 tgtggctgtg gccccagcta acgacatcta caacgagcgc gagctgctga acagcatggg
6001 catcagccag cccaccgtcg tattcgtgag caagaaaggg ctgcaaaaga tcctcaacgt
6061 gcaaaagaag ctaccgatca tacaaaagat catcatcatg gatagcaaga ccgactacca
6121 gggcttccaa agcatgtaca ccttcgtgac ttcccatttg ccacccggct tcaacgagta
6181 cgacttcgtg cccgagagct tcgaccggga caaaaccatc gccctgatca tgaacagtag
6241 tggcagtacc ggattgccca agggcgtagc cctaccgcac cgcaccgctt gtgtccgatt
6301 cagtcatgcc cgcgacccca tcttcggcaa ccagatcatc cccgacaccg ctatcctcag
6361 cgtggtgcca tttcaccacg gcttcggcat gttcaccacg ctgggctact tgatctgcgg
6421 ctttcgggtc gtgctcatgt accgcttcga ggaggagcta ttcttgcgca gcttgcaaga
6481 ctataagatt caatctgccc tgctggtgcc cacactattt agcttcttcg ctaagagcac
6541 tctcatcgac aagtacgacc taagcaactt gcacgagatc gccagcggcg gggcgccgct
6601 cagcaaggag gtaggtgagg ccgtggccaa acgcttccac ctaccaggca tccgccaggg
6661 ctacggcctg acagaaacaa ccagcgccat tctgatcacc cccgaagggg acgacaagcc
6721 tggcgcagta ggcaaggtgg tgcccttctt cgaggctaag gtggtggact tggacaccgg
6781 taagacactg ggtgtgaacc agcgcggcga gctgtgcgtc cgtggcccca tgatcatgag
6841 cggctacgtt aacaaccccg aggctacaaa cgctctcatc gacaaggacg gctggctgca
6901 cagcggcgac atcgcctact gggacgagga cgagcacttc ttcatcgtgg accggctgaa
6961 gagcctgatc aaatacaagg gctaccaggt agccccagcc gaactggaga gcatcctgct
7021 gcaacacccc aacatcttcg acgccggggt cgccggcctg cccgacgacg atgccggcga
7081 gctgcccgcc gcagtcgtcg tgctggaaca cggtaaaacc atgaccgaga aggagatcgt
7141 ggactatgtg gccagccagg ttacaaccgc caagaagctg cgcggtggtg ttgtgttcgt
7201 ggacgaggtg cctaaaggac tgaccggcaa gttggacgcc cgcaagatcc gcgagattct
7261 cattaaggcc aagaagggcg gcaagatcgc cgtgtaattc tagttgttta aacgagctcg
7321 ctagcctcga gtctagagtc gacctgcagg
//

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