Name: efficient sporulation of saccharomyces cerevisiae in a 96 multiwell format
Uploaded: 2016-09-17T14:00:00
Description: Watch this Scientific Journal Video about Efficient Sporulation of Saccharomyces cerevisiae in a 96 Multiwell Format at JoVE.com

وأيد هذا العمل من خلال منحة جوزيف هيلي من جامعة ماساتشوستس في بوسطن (LSH) وR15 GM86805 من المعاهد الوطنية للصحة (LSH). ويدعم SMP جزئيا على زمالة سانوفي-جنزايم في جامعة ماساتشوستس في بوسطن.

1. الاستعداد للتبوغ ملاحظة: وسائل الإعلام وصفها في هذا البروتوكول تتم باستخدام وصفات وطرق معيارية 13،17 يعطي الجدول 1 صياغة ل1 لتر من وسائل الإعلام المختلفة المستخدمة في هذا البروتوكول. <table border="1" cellpadding="0" cellspacing="0" heig...

<ol>
	<li>Marston, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chromosome+segregation+in+budding+yeast:+sister+chromatid+cohesion+and+related+mechanisms.">Chromosome segregation in budding yeast: sister chromatid cohesion and related mechanisms.</a> Genetics. 196 (1), 31-63 (2014).</li><li>Keeney, S., Lange, J., Mohibullah, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Self-organization+of+meiotic+recombination+initiation:+general+principles+and+molecular+pathways.">Self-organization of meiotic recombination initiation: general principles and molecular pathways.</a> Annu. Rev. Genet. 48, 187-214 (2014).</li><li>Granek, J. A., Kayikci, O., Magwene, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pleiotropic+signaling+pathways+orchestrate+yeast+development.">Pleiotropic signaling pathways orchestrate yeast development.</a> Curr. Opin. Microbiol. 14 (6), 676-681 (2011).</li><li>Broach, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nutritional+control+of+growth+and+development+in+yeast.">Nutritional control of growth and development in yeast.</a> Genetics. 192 (1), 73-105 (2012).</li><li>Winter, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Sum1/Ndt80+transcriptional+switch+and+commitment+to+meiosis+in+Saccharomyces+cerevisiae.">The Sum1/Ndt80 transcriptional switch and commitment to meiosis in Saccharomyces cerevisiae.</a> Microbiol. Mol. Biol. Rev. 76 (1), 1-15 (2012).</li><li>Neiman, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sporulation+in+the+budding+yeast+Saccharomyces+cerevisiae.">Sporulation in the budding yeast Saccharomyces cerevisiae.</a> Genetics. 189 (3), 737-765 (2011).</li><li>Padmore, R., Cao, L., Kleckner, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Temporal+comparison+of+recombination+and+synaptonemal+complex+formation+during+meiosis+in+S.+cerevisiae.">Temporal comparison of recombination and synaptonemal complex formation during meiosis in S. cerevisiae.</a> Cell. 66 (6), 1239-1256 (1991).</li><li>Liti, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Population+genomics+of+domestic+and+wild+yeasts.">Population genomics of domestic and wild yeasts.</a> Nature. 458 (7236), 337-341 (2009).</li><li>McCusker, J. H., Haber, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Efficient+sporulation+of+yeast+in+media+buffered+near+pH6.">Efficient sporulation of yeast in media buffered near pH6.</a> J. Bacteriol. 132 (1), 180-185 (1977).</li><li>Codon, A. C., Gasent-Ramirez, J. M., Benitez, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+which+affect+the+frequency+of+sporulation+and+tetrad+formation+in+Saccharomyces+cerevisiae+baker's+yeasts.">Factors which affect the frequency of sporulation and tetrad formation in Saccharomyces cerevisiae baker's yeasts.</a> Appl. Environ. Microbiol. 61 (2), 630-638 (1995).</li><li>Elrod, S. L., Chen, S. M., Schwartz, K., Shuster, E. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimizing+sporulation+conditions+for+different+Saccharomyces+cerevisiae+strain+backgrounds.">Optimizing sporulation conditions for different Saccharomyces cerevisiae strain backgrounds.</a> Methods Mol. Biol. 557, 21-26 (2009).</li><li>B&#246;rner, G. V., Cha, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+yeast+sporulation+efficiency,+spore+viability,+and+meiotic+recombination+on+solid+medium.">Analysis of yeast sporulation efficiency, spore viability, and meiotic recombination on solid medium.</a> Cold Spring Harb. Protoc. 2015 (11), 1003-1008 (2015).</li><li>B&#246;rner, G. V., Cha, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+and+analysis+of+synchronous+meiotic+yeast+cultures.">Induction and analysis of synchronous meiotic yeast cultures.</a> Cold Spring Harb. Protocols. (10), 908-913 (2015).</li><li>Jones, G. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+systematic+library+for+comprehensive+overexpression+screens+in+Saccharomyces+cerevisiae.">A systematic library for comprehensive overexpression screens in Saccharomyces cerevisiae.</a> Nat. Methods. 5 (3), 239-241 (2008).</li><li>Fleming, M. S., Gitler, A. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+yeast+plasmid+overexpession+screen.">High-throughput yeast plasmid overexpession screen.</a> J. Vis. Exp. (53), e2836 (2011).</li><li>Paulissen, S. M., Slubowski, C. J., Roesner, J. M., Huang, L. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Timely+Closure+of+the+Prospore+Membrane+Requires+SPS1+and+SPO77+in+Saccharomyces+cerevisiae.">Timely Closure of the Prospore Membrane Requires SPS1 and SPO77 in Saccharomyces cerevisiae.</a> Genetics. , (2016).</li><li>Amberg, D. C., Burke, D., Strathern, J. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Methods in Yeast Genetics: A Cold Spring Harbor Labooratory Manual. , (2005).</li><li>Parodi, E. M., Baker, C. S., Tetzlaff, C., Villahermosa, S., Huang, L. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SPO71+mediates+prospore+membrane+size+and+maturation+in+Saccharomyces+cerevisiae.">SPO71 mediates prospore membrane size and maturation in Saccharomyces cerevisiae.</a> Eukaryot. Cell. 11 (10), 1191-1200 (2012).</li><li>Nakanishi, H., de Los Santos, P., Neiman, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Positive+and+negative+regulation+of+a+SNARE+protein+by+control+of+intracellular+localization.">Positive and negative regulation of a SNARE protein by control of intracellular localization.</a> Mol. Biol. Cell. 15 (4), 1802-1815 (2004).</li><li>Huang, L. S., Doherty, H. K., Herskowitz, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Smk1p+MAP+kinase+negatively+regulates+Gsc2p,+a+1,3-beta-glucan+synthase,+during+spore+wall+morphogenesis+in+Saccharomyces+cerevisiae.">The Smk1p MAP kinase negatively regulates Gsc2p, a 1,3-beta-glucan synthase, during spore wall morphogenesis in Saccharomyces cerevisiae.</a> Proc. Natl. Acad. Sci. USA. 102 (35), 13431-13436 (2005).</li></ol>

هنا نقدم بروتوكول للsporulating SK1 الخميرة في شكل multiwell 96. التهوية هي المفتاح للتبوغ كفاءة، الأمر الذي يتطلب استخدام إما بقضيب أو حبة الزجاج في كل بئر. عندما يتم sporulated الخلايا في لوحة multiwell 96 في حاضنة تهتز دون أي حبة أو بقضيب، الخلايا لا sporulate بكفاءة. وينظر فقط زيادة صغيرة في ?...

وقد درس تبوغ في الخميرة في مهدها لتقديم رؤى في جوانب كثيرة من الأحياء، بما في ذلك السيطرة على الفصل كروموسوم أثناء الانقسام الاختزالي 1، آليات إعادة التركيب الجيني 2، والسيطرة على التنمية من خلال خلية يشير 3، ومراقبة التغذية التنمية 4، والنسخي تنظيم التنمية 5، والنظر في تشكيل بوغ 6. يتضمن تشكيل بوغ حدث انقسام الخلايا رواية تنطوي على تشكيل المقصورات غشاء جديدة داخل الخلية الأم تليها ترسب جدار بوغ واقية 6. هذه الدراسات أن دراسة الخلايا sporulating في كثير من الأحيان الاستفادة من sporulating بسرعة الخميرة سلالة SK1، والتي يمكن أن تخضع لعملية تكاثر في حوالي 24 ساعة بطريقة فعالة نسبيا 7،8. وعلى الرغم من وصفت تحسين ظروف تبوغ لفي مهدها الخميرة 13/9، هذه التجربةالصورة فحص تبوغ على وسائل الاعلام الصلبة أو في جو من الثقافات السائل على نطاق وحين ينفذ تبوغ من استخدام أنابيب ثقافة أو قوارير. نحن هنا تصف طريقة لsporulating الخميرة في شكل لوحة multiwell 96. نجد أن لهذا الأسلوب، تهوية أمر بالغ الأهمية للتبوغ متزامن وكفاءة، وقد ابتكرت تقنيات لضمان تبوغ كافية لصغيرة الحجم شكل multiwell. Sporulating في شكل 96 لوحة multiwell يسمح للخلايا ليتم عرضه باستخدام تقنيات عالية الإنتاجية والكواشف الأمثل لشكل لوحة multiwell، مثل هذا الفحص لالمكثفات نسخة عالية باستخدام مكتبة مزينة بالبلاط 14-16.

<table border="0" cellpadding="0" cellspacing="0" width="745">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Nunc 1.3 ml DeepWell Plates</td>
			<td style="width:123px;">ThermoScientific</td>
			<td style="width:119px;">260251</td>
			<td style="width:288px;">Used for sporulation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Nunc 2.0 ml DeepWell plates</td>
			<td style="width:123px;">ThermoScientific</td>
			<td>278743</td>
			<td>Used for presporulation growth, step 1.2.3</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">3 mm glass bead</td>
			<td style="width:123px;">Fisher</td>
			<td>11-312A</td>
			<td>Used for sporulation</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">5 mm x 2 mm stir bar, pack of 12</td>
			<td style="width:123px;">Fisher</td>
			<td style="width:119px;">14-511-82</td>
			<td>Used for sporulation</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">96 well frogger</td>
			<td style="width:123px;">V&#38;P Scientific</td>
			<td style="width:119px;">VP407</td>
			<td>needed for step 1.2</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">library copier</td>
			<td style="width:123px;">V&#38;P Scientific</td>
			<td style="width:119px;">VP381</td>
			<td>needed for step 1.2; to be used with the frogger</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">rectangular petri dish</td>
			<td style="width:123px;">ThermoScientific</td>
			<td style="width:119px;">264728</td>
			<td>needed for step 1.2</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Bacto Peptone</td>
			<td style="width:123px;">BD</td>
			<td>211677</td>
			<td>needed for media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Yeast Extract</td>
			<td style="width:123px;">BD</td>
			<td>212750</td>
			<td>needed for media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Bacto Agar</td>
			<td style="width:123px;">BD</td>
			<td style="width:119px;">212750</td>
			<td>needed for media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Dextrose</td>
			<td style="width:123px;">Fisher</td>
			<td style="width:119px;">D16-3</td>
			<td>needed for media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Potassium Acetate</td>
			<td style="width:123px;">Fisher</td>
			<td style="width:119px;">P171-500</td>
			<td>needed for media</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Glycerol</td>
			<td style="width:123px;">Fisher</td>
			<td style="width:119px;">G33-500</td>
			<td>needed for media</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Black 96 well glass bottom plate</td>
			<td style="width:123px;">MatTek</td>
			<td style="width:119px;">PBK96G-1.5.5-F</td>
			<td>needed for step 2.4</td>
		</tr>
	</tbody>
</table>

efficient sporulation of saccharomyces cerevisiae in a 96 multiwell format

During times of nutritional stress, Saccharomyces cerevisiae undergoes gametogenesis, known as sporulation. Diploid yeast cells that are starved for nitrogen and carbon will initiate the sporulation process. The process of sporulation includes meiosis followed by spore formation, where the haploid nuclei are packaged into environmentally resistant spores. We have developed methods for the efficient sporulation of budding yeast in 96 multiwell plates, to increase the throughput of screening yeast cells for sporulation phenotypes. These methods are compatible with screening with yeast containing plasmids requiring nutritional selection, when appropriate minimal media is used, or with screening yeast with genomic alterations, when a rich presporulation regimen is used. We find that for this method, aeration during sporulation is critical for spore formation, and have devised techniques to ensure sufficient aeration that are compatible with the 96 multiwell plate format. Although these methods do not achieve the typical ~80% level of sporulation that can be achieved in large-volume flask based experiments, these methods will reliably achieve about 50-60% level of sporulation in small-volume multiwell plates.

<html> لتقييم هذا البروتوكول، وتمت مقارنة الكفاءة تبوغ تم الحصول عليها من sporulating الخلايا في لوحات multiwell (كما هو موضح أعلاه) إلى خلايا sporulated استخدام كميات أكبر في قوارير (الجدول 2). وقال إن استخدام لوحات multiwell لا يحقق كفاءة عالية ينظر عندما sporulating ?...

Watch this Scientific Journal Video about Efficient Sporulation of Saccharomyces cerevisiae in a 96 Multiwell Format at JoVE.com

Efficient Sporulation of Saccharomyces cerevisiae in a 96 Multiwell Format

Here, sporulation of Saccharomyces cerevisiae is carried out in a 96 multiwell format.

تبوغ الفعال لل<em&gt; خميرة الخباز</em&gt; في تنسيق Multiwell 96

During times of nutritional stress, Saccharomyces cerevisiae undergoes gametogenesis, known as sporulation. Diploid yeast cells that are starved for ...

The overall goal of this protocol is to sporulate budding yeast efficiently in a 96-well format. Although many large-scale screens have been carried out on the budding yeast, high-throughput studies looking at meiosis and sporulation have been limited. The technique described in this video has the advantage of sporulating yeast efficiently in a cost-effective manner in a 96-well format compatible with high-throughput screening.I first got the idea for this method because I wanted to use a tiled two micron plasmid library in order to screen for high copy suppressors of a mutant that could not form refractile spores. Initially, my sporulation efficiencies were poor in the 96-well format. I found this was due to inadequate aeration.Begin this procedure by thawing the SK1 yeast strain onto a yeast extract peptone glycerol or YPG plate. Use a sterile stick to scrape a small amount of cells out of the freezer vial containing the glycerol stock and spread it onto a YPG plate. Grow the yeast for 12 to 16 hours at 30 degrees Celsius.Next, streak the yeast from the YPG plate onto a yeast extract peptone dextrose, or YPD, plate for single colones. Grow the yeast on the YPD plate for 24 to 48 hours at 30 degrees Celsius. After 24 to 48 hours of growth on YPD solid media, start a 25 milliliter YPD culture using a single colony from the YPD plate.Grow this culture overnight in a 30 degree Celsius shaking incubator at 220 RPM until the cells reach an OD600 of 4.0 to 7.0. On the following day, use an appropriate amount of cells from the YPD culture to start a 125 milliliter culture in yeast extract peptone acetate, or YPA, with an OD600 of 0.1. Grow the culture in a 30 degree Celsius shaking incubator at 220 RPM until the OD600 is between 1.3 and 1.5.Transfer the culture to centrifuge tubes and spin at 1800X G in a tabletop centrifuge for five minutes to pellet the cells. Wash the cells once with a 0.5 volume of sterile autoclaved deionized water. Spin again and discard the water.Resuspend the cells in a 1X volume of sporulation media. To prepare cells of different genotypes, cells from thawed frozen stocks in a 96 multiwell plate are first transferred onto YPG solid media. Flame a 96 prong frogger and then cool it momentarily on a plate of solid media before touching it to the yeast colonies.Grow the yeast overnight at 30 degrees Celsius. On the following day, transfer the cells from YPG solid media onto the YPD or selective solid media, depending on the genotype of the strain. YPD is used in this demonstration.Grow the yeast at 30 degrees Celsius until colonies are formed for each strain. Aliquot 1.2 milliliters of YPD liquid media into each well of a 96 deep well plate. Transfer the cells from the solid media into liquid media.Cover the plate with a plastic lid from a rectangular petri plate and hold the lid in place using a piece of tape to minimally restrict air circulation. Grow the cells overnight in a 30 degree Celsius shaker at 220 RPM. On the following day, centrifuge at 1800X G for five minutes.Pour off the media and add 500 microliters of YPA to each well. And resuspend the pelleted cells. Cover the plate with a plastic lid held in place by tape.Grow the cells for 15 hours in a 30 degree Celsius shaking incubator at 220 RPM. To provide proper aeration for efficient sporulation, add into each well of a 96-well plate either one three millimeter sterile solid glass bead or a sterile five millimeter by two millimeter magnetic stir bar. If all cells are of the same genotype, aliquat 500 microliters of cells plus media into each well of the 96-multiwell plate.Cover the plate with a plastic lid held in place by tape. If cells are of different genotypes, spin the cells that are in YPA at 1800X G for five minutes and pour off the media. Add 500 microliters of sporulation media to each well.And resuspend the pelleted cells in the sporulation media. Cover the plate with a plastic lid held in place by tape. If using glass beads, place the samples in a shaking incubator at 30 degrees Celsius and shake at 220 RPM for sporulation.If using magnetic stir bars, place the samples on a stir plate inside of a 30 degrees Celsius incubator for sporulation. The exact stir speed for the magnetic bars is not very important, as long as all of the bars are moving energetically and the culture is aerated. 36 hours later, retrieve the samples to monitor progression through sporulation.Withdraw six microliters of cells from each sporulating culture and add to 24 microliters of sporulation media in a 96-well coverslip plate. Leave the cells to settle for five minutes before examining them using an inverted microscope. A representative sporulation of two different strains is shown in these images of cells visualized using a 63X objective on an inverted microscope.Refractile tetrads, indicated by the arrowheads, can be seen in the wild-type culture. But not in the culture containing the sporulation-deficient smk1 delete cells. Comparisons were made between sporulation efficiencies from cells sporulated in multi-well plates and cells sporulated using larger volumes in flasks.The use of multi-well plates did not achieve the high efficiency routinely seen when using the more typical protocol of sporulating in flasks. Sporulating in multi-well plates with proper aeration can achieve sufficient sporulation efficiencies greater than 50%with the best results obtained using a five millimeter stir bar. However, given the high cost of the small stir bars, the efficiencies obtained using glass beads are comparable.Although both five millimeter and seven millimeter stir bars fit within the well of a 96-multiwell plate, using a seven millimeter stir bar resulted in poor sporulation efficiency. The seven millimeter stir bar in a well tends to interact with a stir bar in an adjacent well. Preventing the bars from stirring properly and providing adequate aeration to the culture.After watching this video, you should have a good understanding of how to efficiently sporulate yeast cells in liquid culture in the 96-well format. Once mastered, this process can be completed in six days, if done properly. This procedure can be used for screening different mutant strains, plasmids, or small molecule inhibitors for their effect on meiosis and sporulation.After it's development, this technique was used to study the regulation of meiotic cell cycle and how cytokinesis and meiosis are coupled in sporulation in Saccharomyces cerevisiae.

Research

JoVE Journal

Biology

Efficient Sporulation of Saccharomyces cerevisiae in a 96 Multiwell Format

Dissection of Saccharomyces Cerevisiae Asci

Dissection of Saccharomyces Cerevisiae Asci

تشريح السكيراء الجعوية زقاق

Yeast is a highly tractable model system that is used to study many different cellular processes. The common laboratory strain Saccharomyces cerevisiae ...

Quantitative Phosphoproteomics in Fatty Acid Stimulated Saccharomyces cerevisiae

Quantitative Phosphoproteomics in Fatty Acid Stimulated Saccharomyces cerevisiae

حفز Phosphoproteomics الكمية في الأحماض الدهنية السكيراء الجعوية</em

This protocol describes the growth and stimulation, with the fatty acid oleate, of isotopically heavy and light S. cerevisiae cells. Cells are ground ...

Microarray Analysis for Saccharomyces cerevisiae

Microarray Analysis for Saccharomyces cerevisiae

تحليل لميكروأري السكيراء الجعوية</em

In this protocol, gene expression in yeast (Saccharomyces cerevisiae) is changed after exposure to oxidative stress induced by the addition of hydrogen ...

A Fluorescence Microscopy Assay for Monitoring Mitophagy in the Yeast Saccharomyces cerevisiae

A Fluorescence Microscopy Assay for Monitoring Mitophagy in the Yeast Saccharomyces cerevisiae

والفحص المجهري لالإسفار Mitophagy الرصد في الخميرة السكيراء الجعوية

Autophagy is important for turnover of cellular components under a range of different conditions. It serves an essential homeostatic function as well as a ...

Rapid and Efficient Generation of Neurons from Human Pluripotent Stem Cells in a Multititre Plate Format

بسرعة وكفاءة توليد الخلايا العصبية من الخلايا الجذعية المحفزة الإنسان في تنسيق اللوحة Multititre

Existing protocols for the generation of neurons from human pluripotent stem cells (hPSCs) are often tedious in that they are  multistep procedures ...

Visualization and Analysis of mRNA Molecules Using Fluorescence In Situ Hybridization in Saccharomyces cerevisiae

Visualization and Analysis of mRNA Molecules Using Fluorescence In Situ Hybridization in Saccharomyces cerevisiae

التصور والتحليل من جزيئات الرنا المرسال عن طريق الإسفار<em&gt; في الموضع</em&gt; التهجين في<em&gt; خميرة الخباز</em

The Fluorescence in situ Hybridization (FISH) method allows one to detect nucleic acids in the native cellular environment. Here we provide a protocol for ...

Coupled Assays for Monitoring Protein Refolding in Saccharomyces cerevisiae

Coupled Assays for Monitoring Protein Refolding in Saccharomyces cerevisiae

فحوصات بالإضافة لرصد طوي ثانية بروتين في<em&gt; خميرة الخباز</em

Proteostasis, defined as the combined processes of protein folding/biogenesis, refolding/repair, and degradation, is a delicate cellular balance that must ...

Purification of the Cystic Fibrosis Transmembrane Conductance Regulator Protein Expressed in Saccharomyces cerevisiae

Purification of the Cystic Fibrosis Transmembrane Conductance Regulator Protein Expressed in Saccharomyces cerevisiae

تنقية التليف الكيسي عبر الغشاء المواصلة منظم البروتين المعبر عنه في<em&gt; خميرة الخباز</em

Defects in the cystic fibrosis transmembrane conductance regulator (CFTR) protein cause cystic fibrosis (CF), an autosomal recessive disease that ...

Measurement of mRNA Decay Rates in Saccharomyces cerevisiae Using rpb1-1 Strains

Measurement of mRNA Decay Rates in Saccharomyces cerevisiae Using rpb1-1 Strains

قياس مرنا تسوس الأسعار<em&gt; خميرة الخباز</em&gt; استخدام<em&gt; rpb1-1</em&gt; سلالات

mRNA steady state levels vary depending on environmental conditions. Regulation of the steady state accumulation levels of an mRNA ensures that the ...