Name: studying age-dependent genomic instability using the s. cerevisiae chronological lifespan model
Uploaded: 2011-09-29T12:00:00
Duration: 8 min 46 s
Description: Watch this Scientific Journal Video about Studying Age-dependent Genomic Instability using the S. cerevisiae Chronological Lifespan Model at JoVE.com

我々は、プラスミド、酵母菌株を提供するためのS.ジンクスロバートソンとE ·ハイデンリッヒに感謝、P. PhamさんとMFグッドマンヘルプwiththe TLSアッセイのために。この作品は、エイジング総合研究助成のためのアメリカ連合によっておよびNIH AG20642、AG025135によって、部分的に、サポートされていました。

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液体老化の文化はいつか再生/ CLS分析を複雑にあえぎ表現型13を 、示す。人口以上の90から99パーセントが生存能力を失ったときに再生が一般的に発生します。この表現型は、多くの場合、酸化ストレスおよび/または細胞の減少の保護に関連付けられています。例えば、この表現型の頻度は、より多くの細胞質スーパーオキシドジスムターゼを欠く細胞で2倍以上と大幅に長寿命変異体で減少します（例えば、sch9ΔまたはRAS2Δ）または変異体を過剰発現してスーパーオキシドは、22ジスムターゼ。実際には、我々は、経時的老化時の高死亡率の段階で3つの連続した​​サンプル用実行可能性の実現可能性または安定化の増加として再成長を定義する。 異なるDNAの変異の年齢に依存する周波数が大幅にひずみ背景、遺伝子操作、培養条件、ならびに再生/あえぎと非常に低い生存率に応じて異なります。プレ実験はこのようなperformingtheフルスケールの寿命解析の前に変異の周波数範囲を決定するために突然変異試験のための様々な播種密度と平均値と最大生存などの時点で行ってください。野生型の菌株は、常に、任意の治療や遺伝的変異体と平行に寿命や突然変異の頻度の研究に含まれるべきである間実験-ばらつきが考慮できるように。複数の生物学的なレプリカが研究に含まれるべき、と、液体の両方とその場可能性/突然変異アッセイの結果を裏付けるために実施すべきである。 代わりに組み合わせて複数のアッセイを用いて年齢依存性ゲノム不安定性のプロファイリング、DNAの突然変異の特定のタイプに焦点を当ててから、年齢依存性のゲノム不安定性に寄与する特定のDNA損傷とDNA損傷修復システム（秒）に光を当てることができる。例えば、他のDNAの突然変異のものに比べて総染色体再配列（GCRs）の有意な増加が、、elevationof二重鎖切断および/または酵母中の接合端（NHEJ）非相同の減損を示唆する野生型酵母で観察される経時的老化（表2）。野生型エージング酵母で得られたCAN1突然変異のスペクトルは、（CAN1遺伝子の塩基配列決定）経時的老化とエラーが発生しやすいDNAの修復中に酸化的損傷の増加を示唆し、一方、長寿命のsch9Δ変異体では、以下の酸化的DNA損傷と程度に下げてエラーを起こしやすいtranslesionの合成は、20を観察した。 ここで説明する方法は、さらに年齢依存性のゲノム不安定性を研究するために費やさすることができます。例えば、静止及び非静止期の細胞は、Allenらによって記述された密度勾配法を用いて酵母固定相培養細胞から分離することができます。23。ここで説明する変異のアッセイと組み合わせることで、我々は、年齢依存性の変異の大部分は静止細胞ではなく、分割し、損傷やアポトーシス細胞20,24から生じることが報告されている。

studying age-dependent genomic instability using the s. cerevisiae chronological lifespan model

Saccharomyces cerevisiaeの老化モデルを用いた研究は、部分的に高等真核生物1-2に保存されている寿命調節経路を明らかにしました。酵母老化モデルのシンプルさとパワーは、DNA損傷とゲノム維持だけでなく、老化の間に病気への貢献を研究するために調べることができます。ここで、我々は単純なDNAで酵母の時系列の寿命を組み合わせることにより、塩基置換、フレームシフト変異、総染色体再配列、および相同/ homeologous組換えだけでなく、核DNA修復活性を含め、年齢依存性のDNA変異を研究するためにシステムを説明損傷と突然変異試験。ここで説明する方法は、ゲノムの不安定性と哺乳類では年齢依存性DNAの変異と癌の根底にあるメカニズムを制御する遺伝子/経路の同定を容易にしてください。

Watch this Scientific Journal Video about Studying Age-dependent Genomic Instability using the S. cerevisiae Chronological Lifespan Model at JoVE.com

Studying Age-dependent Genomic Instability using the S. cerevisiae Chronological Lifespan Model

ここでは、エージング中規制またはゲノムDNAの不安定性に寄与する遺伝子/経路を研究するために酵母の時系列の寿命モデルと組み合わせることができるDNAの突然変異試験のセットを説明します。

使用年齢に依存するゲノム不安定性を研究して S.出芽酵母時系列寿命モデル

Studies using the Saccharomyces cerevisiae aging model have uncovered life span regulatory pathways that are partially conserved in higher eukaryotes1-2. ...

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Biology

Studying Age-dependent Genomic Instability using the S. cerevisiae Chronological Lifespan Model

Assay for Adhesion and Agar Invasion in S. cerevisiae

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Yeasts are found in natural biofilms, where many microorganisms colonize surfaces. In artificial environments, such as surfaces of man-made objects, ...

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High-throughput methods to examine protein localization or organelle morphology is an effective tool for studying protein interactions and can help ...

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Lipids are the building blocks of cellular membranes that function as barriers and in compartmentalization of cellular processes, and recently, as ...

Microarray Analysis for Saccharomyces cerevisiae

Microarray Analysis for Saccharomyces cerevisiae

マイクロアレイ解析のためのサッカロマイセスセレビシエ</em

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

Genomic MRI - a Public Resource for Studying Sequence Patterns within Genomic DNA

ゲノムMRI - ゲノムDNA内の配列パターンを研究するためのパブリックリソース

Non-coding genomic regions in complex eukaryotes, including intergenic areas, introns, and untranslated segments of exons, are profoundly non-random in ...

Flow Cytometry-based Purification of S. cerevisiae Zygotes

Flow Cytometry-based Purification of S. cerevisiae Zygotes

のフローサイトメトリーベースの精製<em&gt; S.セレビ</em&gt;受精卵

Zygotes are essential intermediates between haploid and diploid states in the life cycle of many organisms, including yeast (Figure 1) 1. S. cerevisiae ...

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蛍光クリアランスアッセイを用いてゼブラフィッシュ腎臓機能の評価

The zebrafish embryo offers a tractable model to study organogenesis and model human genetic disease. Despite its relative simplicity, the zebrafish ...

Using Microfluidic Devices to Measure Lifespan and Cellular Phenotypes in Single Budding Yeast Cells

シングル出芽酵母細胞での寿命と細胞表現型を測定するためのマイクロ流体デバイスを使用しました

Budding yeast Saccharomyces cerevisiae is an important model organism in aging research. Genetic studies have revealed many genes with conserved effects ...

High-throughput Screening for Protein-based Inheritance in S. cerevisiae

High-throughput Screening for Protein-based Inheritance in S. cerevisiae

出芽酵母におけるタンパク質ベース継承の高速スクリーニング

The encoding of biological information that is accessible to future generations is generally achieved via changes to the DNA sequence. Long-lived ...

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腰椎不安定性のマウスモデル

Intervertebral disc degeneration (IDD) is a common pathological change leading to low back pain. Appropriate animal models are desired for understanding ...