Name: Fare embriyonik kök hücrelerinde Enhancer RNA Stabilitesinin Optimize Edilebilir Nicel Değerlendirmesi (Video) | JoVE Journal
Uploaded: 2025-11-21T14:45:00.000+00:00
Duration: 3 min 34 s
Description: Keşfedin fare embriyonik kök hücrelerinde enhancer RNA stabilitesinin nicel değerlendirmesini. RT-qPCR ve doğrusal olmayan regresyon analizi kullanarak eRNA yarı ömrünü ve bozunma dinamiklerini doğru bir şekilde modelleyin.

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<ol><li>Bulger, M., Groudine, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Functional+and+mechanistic+diversity+of+distal+transcription+enhancers%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Functional and mechanistic diversity of distal transcription enhancers.</a> Cell. 144 (3), 327-339 (2011).</li>
<li>Gorbovytska, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhancer+RNAs+stimulate+Pol+II+pause+release+by+harnessing+multivalent+interactions+to+NELF%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Enhancer RNAs stimulate Pol II pause release by harnessing multivalent interactions to NELF.</a> Nat Commun. 13 (1), 2429(2022).</li>
<li>Joo, J. -Y., Schaukowitch, K., Farbiak, L., Kilaru, G., Kim, T. -K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Stimulus-specific+combinatorial+functionality+of+neuronal+c-fos+enhancers%5BTitle%5D)+AND+%22Nat+Neurosci%22%5BJournal%5D)">Stimulus-specific combinatorial functionality of neuronal c-fos enhancers.</a> Nat Neurosci. 19 (1), 75-83 (2016).</li>
<li>Levine, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Transcriptional+enhancers+in+animal+development+anfd+evolution%5BTitle%5D)+AND+%22Curr+Biol%22%5BJournal%5D)">Transcriptional enhancers in animal development anfd evolution.</a> Curr Biol. 20 (17), R754-R763 (2010).</li>
<li>Nord, A. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Rapid+and+pervasive+changes+in+genome-wide+enhancer+usage+during+mammalian+development%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Rapid and pervasive changes in genome-wide enhancer usage during mammalian development.</a> Cell. 155 (7), 1521-1531 (2013).</li>
<li>Schwalb, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((TT-seq+maps+the+human+transient+transcriptome%5BTitle%5D)+AND+%22Science%22%5BJournal%5D)">TT-seq maps the human transient transcriptome.</a> Science. 352 (6290), 1225-1228 (2016).</li>
<li>De Santa, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+large+fraction+of+extragenic+RNA+pol+II+transcription+sites+overlap+enhancers%5BTitle%5D)+AND+%22PLoS+Biol%22%5BJournal%5D)">A large fraction of extragenic RNA pol II transcription sites overlap enhancers.</a> PLoS Biol. 8 (5), e1000384(2010).</li>
<li>Kim, T. -K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Widespread+transcription+at+neuronal+activity-regulated+enhancers%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Widespread transcription at neuronal activity-regulated enhancers.</a> Nature. 465 (7295), 182-187 (2010).</li>
<li>Songyue, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+roles+of+enhancer%2C+especially+super-enhancer-driven+genes+in+tumor+metabolism+and+immunity%5BTitle%5D)+AND+%22Int+J+Biol+Macromol%22%5BJournal%5D)">The roles of enhancer, especially super-enhancer-driven genes in tumor metabolism and immunity.</a> Int J Biol Macromol. 308 (1), 142414(2025).</li>
<li>Kim, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Epigenetic+regulation+of+IFITM1+expression+in+lipopolysaccharide-stimulated+human+mesenchymal+stromal+cells%5BTitle%5D)+AND+%22Stem+Cell+Res+Ther%22%5BJournal%5D)">Epigenetic regulation of IFITM1 expression in lipopolysaccharide-stimulated human mesenchymal stromal cells.</a> Stem Cell Res Ther. 11, 1-12 (2020).</li>
<li>Li, W., Notani, D., Rosenfeld, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhancers+as+non-coding+RNA+transcription+units%3A+Recent+insights+and+future+perspectives%5BTitle%5D)+AND+%22Nat+Rev+Genet%22%5BJournal%5D)">Enhancers as non-coding RNA transcription units: Recent insights and future perspectives.</a> Nat Rev Genet. 17 (4), 207-223 (2016).</li>
<li>Sartorelli, V., Lauberth, S. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhancer+RNAs+are+an+important+regulatory+layer+of+the+epigenome%5BTitle%5D)+AND+%22Nat+Struct+Mol+Biol%22%5BJournal%5D)">Enhancer RNAs are an important regulatory layer of the epigenome.</a> Nat Struct Mol Biol. 27 (6), 521-528 (2020).</li>
<li>Lam, M. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Rev-Erbs+repress+macrophage+gene+expression+by+inhibiting+enhancer-directed+transcription%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Rev-Erbs repress macrophage gene expression by inhibiting enhancer-directed transcription.</a> Nature. 498 (7455), 511-515 (2013).</li>
<li>Li, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Functional+roles+of+enhancer+RNAs+for+oestrogen-dependent+transcriptional+activation%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Functional roles of enhancer RNAs for oestrogen-dependent transcriptional activation.</a> Nature. 498 (7455), 516-520 (2013).</li>
<li>Melo, C. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((eRNAs+are+required+for+p53-dependent+enhancer+activity+and+gene+transcription%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">eRNAs are required for p53-dependent enhancer activity and gene transcription.</a> Mol Cell. 49 (3), 524-535 (2013).</li>
<li>Mousavi, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((eRNAs+promote+transcription+by+establishing+chromatin+accessibility+at+defined+genomic+loci%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">eRNAs promote transcription by establishing chromatin accessibility at defined genomic loci.</a> Mol Cell. 51 (5), 606-617 (2013).</li>
<li>Schaukowitch, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhancer+RNA+facilitates+NELF+release+from+immediate+early+genes%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">Enhancer RNA facilitates NELF release from immediate early genes.</a> Mol Cell. 56 (1), 29-42 (2014).</li>
<li>Göttgens, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Establishing+the+transcriptional+programme+for+blood%3A+The+SCL+stem+cell+enhancer+is+regulated+by+a+multiprotein+complex+containing+Ets+and+GATA+factors%5BTitle%5D)+AND+%22The+EMBO+J%22%5BJournal%5D)">Establishing the transcriptional programme for blood: The SCL stem cell enhancer is regulated by a multiprotein complex containing Ets and GATA factors.</a> The EMBO J. 21 (12), 3039-3050 (2002).</li>
<li>Wang, H. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Defining+essential+enhancers+for+pluripotent+stem+cells+using+a+features-oriented+CRISPR-Cas9+screen%5BTitle%5D)+AND+%22Cell+Rep%22%5BJournal%5D)">Defining essential enhancers for pluripotent stem cells using a features-oriented CRISPR-Cas9 screen.</a> Cell Rep. 33 (4), 108309(2020).</li>
<li>Cinghu, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Intragenic+enhancers+attenuate+host+gene+expression%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">Intragenic enhancers attenuate host gene expression.</a> Mol Cell. 68 (1), 104-117.e106 (2017).</li>
<li>Hobson, D. J., Wei, W., Steinmetz, L. M., Svejstrup, J. Q. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((RNA+polymerase+II+collision+interrupts+convergent+transcription%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">RNA polymerase II collision interrupts convergent transcription.</a> Mol Cell. 48 (3), 365-374 (2012).</li>
<li>Moon, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Embryonic+stem+cell-specific+intragenic+enhancer+RNA+essential+for+NSUN2-mediated+stem+cell+fate+regulation%5BTitle%5D)+AND+%22Int+J+Biol+Macromol%22%5BJournal%5D)">Embryonic stem cell-specific intragenic enhancer RNA essential for NSUN2-mediated stem cell fate regulation.</a> Int J Biol Macromol. 319, 145470(2025).</li>
<li>Tata, P. R., Tata, N. R., Kühl, M., Sirbu, I. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Identification+of+a+novel+epigenetic+regulatory+region+within+the+pluripotency+associated+microRNA+cluster%2C+EEmiRC%5BTitle%5D)+AND+%22Nucleic+Acids+Res%22%5BJournal%5D)">Identification of a novel epigenetic regulatory region within the pluripotency associated microRNA cluster, EEmiRC.</a> Nucleic Acids Res. 39 (9), 3574-3581 (2011).</li>
<li>Sohni, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Dynamic+switching+of+active+promoter+and+enhancer+domains+regulates+Tet1+and+Tet2+expression+during+cell+state+transitions+between+pluripotency+and+differentiation%5BTitle%5D)+AND+%22Mol+Cell+Biol%22%5BJournal%5D)">Dynamic switching of active promoter and enhancer domains regulates Tet1 and Tet2 expression during cell state transitions between pluripotency and differentiation.</a> Mol Cell Biol. 35 (6), 1026-1042 (2015).</li>
<li>Borsari, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enhancers+with+tissue-specific+activity+are+enriched+in+intronic+regions%5BTitle%5D)+AND+%22Genome+Res%22%5BJournal%5D)">Enhancers with tissue-specific activity are enriched in intronic regions.</a> Genome Res. 31 (8), 1325-1336 (2021).</li>
<li>Bressin, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((High-sensitive+nascent+transcript+sequencing+reveals+BRD4-specific+control+of+widespread+enhancer+and+target+gene+transcription%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">High-sensitive nascent transcript sequencing reveals BRD4-specific control of widespread enhancer and target gene transcription.</a> Nat Commun. 14 (1), 4971(2023).</li>
<li>Heintzman, N. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Distinct+and+predictive+chromatin+signatures+of+transcriptional+promoters+and+enhancers+in+the+human+genome%5BTitle%5D)+AND+%22Nat+Genet%22%5BJournal%5D)">Distinct and predictive chromatin signatures of transcriptional promoters and enhancers in the human genome.</a> Nat Genet. 39 (3), 311-318 (2007).</li>
<li>Pérez-Molina, R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((An+intronic+Alu+element+attenuates+the+transcription+of+a+long+non-coding+RNA+in+human+cell+lines%5BTitle%5D)+AND+%22Front+Genet%22%5BJournal%5D)">An intronic Alu element attenuates the transcription of a long non-coding RNA in human cell lines.</a> Front Genet. 11, 928(2020).</li>
<li>Djebali, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Landscape+of+transcription+in+human+cells%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Landscape of transcription in human cells.</a> Nature. 489 (7414), 101-108 (2012).</li>
<li>Rada-Iglesias, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+unique+chromatin+signature+uncovers+early+developmental+enhancers+in+humans%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">A unique chromatin signature uncovers early developmental enhancers in humans.</a> Nature. 470 (7333), 279-283 (2011).</li>
<li>Atlasi, Y., Stunnenberg, H. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+interplay+of+epigenetic+marks+during+stem+cell+differentiation+and+development%5BTitle%5D)+AND+%22Nat+Rev+Genet%22%5BJournal%5D)">The interplay of epigenetic marks during stem cell differentiation and development.</a> Nat Rev Genet. 18 (11), 643-658 (2017).</li>
<li>Young, R. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Control+of+the+embryonic+stem+cell+state%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Control of the embryonic stem cell state.</a> Cell. 144 (6), 940-954 (2011).</li>
<li>Ratnadiwakara, M., Änkö, M. -L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((mRNA+stability+assay+using+transcription+inhibition+by+actinomycin+D+in+mouse+pluripotent+stem+cells%5BTitle%5D)+AND+%22Bio+Protoc%22%5BJournal%5D)">mRNA stability assay using transcription inhibition by actinomycin D in mouse pluripotent stem cells.</a> Bio Protoc. 8 (21), e3072-e3072 (2018).</li>
<li>Kim, S. -K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Functional+coordination+of+BET+family+proteins+underlies+altered+transcription+associated+with+memory+impairment+in+fragile+X+syndrome%5BTitle%5D)+AND+%22Sci+Adv%22%5BJournal%5D)">Functional coordination of BET family proteins underlies altered transcription associated with memory impairment in fragile X syndrome.</a> Sci Adv. 7 (21), eabf7346(2021).</li>
<li>Peterson, S. M., Freeman, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((RNA+Isolation+from+Embryonic+Zebrafish+and+cDNA+Synthesis+for+Gene+Expression+Analysis%5BTitle%5D)+AND+%22J+Vis+Exp%22%5BJournal%5D)">RNA Isolation from Embryonic Zebrafish and cDNA Synthesis for Gene Expression Analysis.</a> J Vis Exp. (30), e1470(2009).</li>
<li>Ye, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Primer-BLAST%3A+A+tool+to+design+target-specific+primers+for+polymerase+chain+reaction%5BTitle%5D)+AND+%22BMC+Bioinformatics%22%5BJournal%5D)">Primer-BLAST: A tool to design target-specific primers for polymerase chain reaction.</a> BMC Bioinformatics. 13 (1), 134(2012).</li>
<li>Kim, S. -K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((SET7%2F9+methylation+of+the+pluripotency+factor+LIN28A+is+a+nucleolar+localization+mechanism+that+blocks+let-7+biogenesis+in+human+ESCs%5BTitle%5D)+AND+%22Cell+Stem+Cell%22%5BJournal%5D)">SET7/9 methylation of the pluripotency factor LIN28A is a nucleolar localization mechanism that blocks let-7 biogenesis in human ESCs.</a> Cell Stem Cell. 15 (6), 735-749 (2014).</li>
<li>Lugowski, A., Nicholson, B., Rissland, O. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((DRUID%3A+A+pipeline+for+transcriptome-wide+measurements+of+mRNA+stability%5BTitle%5D)+AND+%22RNA%22%5BJournal%5D)">DRUID: A pipeline for transcriptome-wide measurements of mRNA stability.</a> RNA. 24 (5), 623-632 (2018).</li>
<li>Lugowski, A., Nicholson, B., Rissland, O. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Determining+mRNA+half-lives+on+a+transcriptome-wide+scale%5BTitle%5D)+AND+%22Methods%22%5BJournal%5D)">Determining mRNA half-lives on a transcriptome-wide scale.</a> Methods. 137, 90-98 (2018).</li>
<li>Tsai, P. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+muscle-specific+enhancer+rna+mediates+cohesin+recruitment+and+regulates+transcription+in+trans%5BTitle%5D)+AND+%22Mol+Cell%22%5BJournal%5D)">A muscle-specific enhancer rna mediates cohesin recruitment and regulates transcription in trans.</a> Mol Cell. 71 (1), 129-141.e128 (2018).</li>
<li>Woolley, C. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Full-length+mRNA+sequencing+resolves+novel+variation+in+5%27+UTR+length+for+genes+expressed+during+human+CD4+T-cell+activation%5BTitle%5D)+AND+%22Immunogenetics%22%5BJournal%5D)">Full-length mRNA sequencing resolves novel variation in 5' UTR length for genes expressed during human CD4 T-cell activation.</a> Immunogenetics. 77 (1), 14(2025).</li>
<li>Zhang, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Algorithm+for+optimized+mRNA+design+improves+stability+and+immunogenicity%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Algorithm for optimized mRNA design improves stability and immunogenicity.</a> Nature. 621 (7978), 396-403 (2023).</li>
<li>Kim, T. -K., Shiekhattar, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Architectural+and+functional+commonalities+between+enhancers+and+promoters%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Architectural and functional commonalities between enhancers and promoters.</a> Cell. 162 (5), 948-959 (2015).</li>
</ol>

<table><tbody><tr><td>2-merkaptoetanol</td><td>Gibco</td><td>21985023</td><td></td></tr><tr><td>Aktinomisin D</td><td>Sigma-Aldrich</td><td>A9415</td><td></td></tr><tr><td>kloroform</td><td>Daejung kimyasalları</td><td>2548-4105</td><td></td></tr><tr><td>DMSO</td><td>GenDEPOT</td><td>D2680</td><td></td></tr><tr><td>Fetal sığır serumu (FBS)</td><td>Sigma-Aldrich</td><td>F0392</td><td></td></tr><tr><td>Jelatin</td><td>Sigma-Aldrich</td><td>G1890-100G</td><td></td></tr><tr><td>Glasgow' Minimum Temel Ortam (GMEM)</td><td>Gibco</td><td>11710035</td><td></td></tr><tr><td>GraphPad Prism (sürüm 9)</td><td>GraphPad Yazılımı</td><td></td><td></td></tr><tr><td>izopropanol</td><td>Daejung kimyasalları</td><td>5035-4105</td><td></td></tr><tr><td>KnockOut Serum Değişimi (KOSR)</td><td>Gibco,&nbsp;</td><td>10828028</td><td></td></tr><tr><td>Lösemi inhibitör faktörü (LIF)</td><td>Merck</td><td>ESG1106</td><td></td></tr><tr><td>MEM Temel Olmayan Amino Asitler Çözeltisi (MnEA)</td><td>Welgene</td><td>LS005</td><td></td></tr><tr><td>M-MLV cDNA Sentez Kiti&nbsp;</td><td>Enzimomikler</td><td>EZ006S</td><td></td></tr><tr><td>OPTIZEN NanoQ Plus&nbsp;</td><td>KLAB</td><td>01-QI685-M-16-C</td><td></td></tr><tr><td>Penisillin-streptomisin</td><td>Welgene</td><td>LS202-02</td><td></td></tr><tr><td>Power SYBR-Green PCR master mix&nbsp;</td><td>Uygulamalı Biyosistemler</td><td>4368708</td><td></td></tr><tr><td>QuanStudio1 Gerçek Zamanlı PCR sistemi&nbsp;</td><td>Uygulamalı Biyosistemler</td><td>A40425</td><td></td></tr><tr><td>Sodyum piruvat</td><td>Welgene</td><td>LS013</td><td></td></tr><tr><td>TRIzol&nbsp;</td><td>Invitrogen</td><td>15596018</td><td></td></tr></tbody></table>

fare embriyonik kök hücrelerinde enhancer rna stabilitesinin optimize edilebilir nicel değerlendirmesi

Optimized Quantitative Assessment of Enhancer RNA Stability in Mouse Embryonic Stem Cells

Fare embriyonik kök hücrelerinde Enhancer RNA Stabilitesinin Optimize Edilebilir Nicel Değerlendirmesi

Enhancer RNAs (eRNAs) are non-coding transcripts produced from transcriptionally active enhancer regions and are increasingly recognized as key regulators ...

optimized-quantitative-assessment-enhancer-rna-stability-mouse

Research

JoVE Journal

Biochemistry

Biyokimya

Biz hızla belirlemek ve fare embriyonik kök hücre bakım ve kendini yenileme düzenleyen genlerin karakterize bir floresan reporter assay açıklar.

Fare Embriyonik Kök Hücre Bakım ve Kendini yenileme Regulate Genler Okumak Oct4GiP Reporter Assay

Tek hücre gen sentezleme tahlili kök hücre heterojeniteler anlamak için gereklidir.

Kantitatif RT-PCR ile Bireysel İnsan Embriyonik Kök Hücreleri profilleme

Geliştirici aktivitenin deneysel doğrulamasına en iyi fonksiyon kaybı analizi ile yaklaşılır. Burada, hibrit bir genom (Mus musculus129 x Mus castaneus) içeren F1 ES hücrelerinde gen transkripsiyonunun alele özgü regülasyonunu incelemek için CRISPR/Cas9 aracılı delesyon kullanan etkili bir protokol sunulmaktadır.

Yaratma CRISPR / Cas9 aracılığı monoallelik Delesyonlar Fare Embriyonik Kök Hücreleri Artırıcı Fonksiyonu Eğitim için

Bu protokol Fare preimplantasyon embriyolar soy özellikleri çalışma protein ekspresyonu in situ analiz kantitatif, tek hücreli gerçekleştirmek için bir yöntem sunmaktadır. blastosist toplanması için gerekli işlemler, proteinlerin immunofluorescent algılama tüm montaj, bir konfokal mikroskop ve nükleer segmentasyon ve görüntü analizi örneklerin görüntüleme açıklanmıştır.

Protein İfade Kantitatif Analiz Fare Preimplantasyon Embriyolar Lineage Şartname Eğitim için

Bu çalışmada, aktif geliştiricilerin nasıl tanımlanabileceğine ve deneysel olarak doğrulanabileceğine dair deneysel bir iş akışı sağlıyoruz.

Tahmin ve Gen Düzenleme Elementlerin Doğrulama Retinoik Asit Kaynaklı Embriyonik sırasında Aktif Hücre Farklılaşma Kök

Proteinler genellikle farklı hücresel fonksiyonları sarf edebilirsiniz birden çok etki içerirler. Gen irtibat kutusu kapağı (KO) bu işlevsel çeşitliliği dikkate almaz. Burada, çeşitli işlevsel etki alanları veya bir proteinin varyantlarının moleküler diseksiyon sağlar KO embriyonik kök hücrelerin bir rekombinasyon aracılı kaset değişimi (RMCE) tabanlı bir yapı-işlev yaklaşım sunulmuştur.

Recombinase aracılı Kaset Değişimi Kullanarak Fare Embriyonik Kök Hücre yapı-fonksiyon Çalışmaları

Bu iletişim kuralını tasarlayın ve çoğaltılmış arttırıcılar deactivating füzyon protein SID4X-dCas9-KRAB, olarak da bilinen artırıcı girişim (Enhancer-i) ile hedefleme gerçekleştirmek için gereken adımları açıklar. Bu iletişim kuralı gen ekspresyonu düzenleyen arttırıcılar tanımlaması sağlar ve ortak bir hedef geni düzenleyen arttırıcılar arasındaki ilişkileri diseksiyon kolaylaştırır.

Diseksiyon artırıcı işlevi hücre hatlarında Multiplex artırıcı CRISPR tabanlı girişim kullanma

Embriyonik kök hücrelerden üretilen embriyoid cisimlerde yüksek iş eldeli sıralamanın ardından kantitatif kromozom konformasyon yakalama uygulamasını rapor ediyoruz. Bu teknik, embriyonik kök hücre farklılaşması sırasında belirli bir genin putatif arttırıcılar ve promotör bölgeleri arasındaki temasları tanımlamaya ve quantitate sağlar.

Kantitatif Kromozom Konformasyon Yakalama (4C) ile Embriyoid Cisimlerde Arttırıcı-Organizatör Kontalarının Belirlenmesi

Bu protokol, yeni bir rAAV tabanlı geçici arttırıcı-muhabir analizini açıklamaktadır. Bu tahlil, fare beyninde in vivo olarak arttırıcı güdümlü ifadeyi indüklemek için kullanılabilir.

Geliştirici Tabanlı İfade Yapılarının AAV Dağıtımı In Vivo in Mouse Brain

Bu protokol, yeni ortaya çıkan geliştirici transkriptleri ölçmek için kolaylaştırılmış bir hesaplama hattı sağlar. Kromatin erişilebilirliğini, kromatin özelliğini ve transkripsiyonel verileri entegre ederek, karmaşık intragenik bölgelerdeki güçlendirici aktivitenin doğru tespitine ve ipliğe özgü analizine olanak tanırken, kapsamlı biyoinformatik eğitimi olmayan araştırmacılar için de erişilebilir olmaya devam eder.

Fare Embriyonik Kök Hücrelerinde İntergenik/İntojenik Güçlendirici RNA Miktar Tayini için Hesaplamalı Bir Boru Hattı

Keşfedin fare embriyonik kök hücrelerinde enhancer RNA stabilitesinin nicel değerlendirmesini. RT-qPCR ve doğrusal olmayan regresyon analizi kullanarak eRNA yarı ömrünü ve bozunma dinamiklerini doğru bir şekilde modelleyin.

Fare embriyonik kök hücrelerinde Enhancer RNA Stabilitesinin Optimize Edilebilir Nicel Değerlendirmesi

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