Understanding YACs and Chromatin Formation

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Introduction to YACs and Chromatin Formation

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Yeast Artificial Chromosomes (YACs) play a critical role in the formation of chromatin within the nucleus of yeast cells. This article aims to explore the relationship between YACs and chromatin, including their replication and condensation processes. We will also delve into the specific characteristics of YACs and their relevance in the context of chromatin biology.

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YACs within the Mitotic Cycle

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Behavior of YACs in Vitro and In Vivo

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YACs, when isolated in vitro (outside the cell), behave as bare or unduplicated DNA sequences. However, inside the yeast nucleus, YACs exhibit behaviors similar to native yeast chromosomes. They recruit histones for repair and occasionally get transcribed into mRNA. It's important to note that while histones are key components of chromatin structure, they are not the primary drivers of chromosome condensation during mitosis.

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Chromatin Structure and Condensation

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Roles of Condensins in Chromosome Condensation

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Metaphase chromosomes, which are highly condensed, are packed with condensins. Condensins are not present on interphase chromosomes, which are less condensed. This difference in condensation patterns during the cell cycle is crucial for the orderly progression of mitosis.

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YACs: Linear or Circular

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Characteristics of YACs

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YACs can be either linear or circular. Circular YACs are more commonly used because they can be easily separated from the linear chromosomes of the host yeast. They contain a yeast centromere, which attaches to the mitotic spindle during telophase, ensuring the accurate distribution of genetic material to daughter cells.

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Replication of YACs

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ARS Sequences and DNA Replication

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YACs are replicated based on the presence of ARS (Autonomously Replicating Sequences) that mark specific DNA regions. These regions are recognized by ORC (Origin Recognition Complex), which initiates DNA replication. The ARS motif (5- T/A T T T A Y R T T T T/A -3) is frequently found in the human genome, allowing for efficient replication in yeast. This characteristic enables the use of YACs in yeast for generating genetically engineered cells.

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YACs and Human Gene Expression

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Expression of Foreign Genes in Yeast

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Unlike Human Artificial Chromosomes (HACs), YACs are not used to express foreign genes in yeast. Human transcription factors, which are essential for gene expression in human cells, are not present in yeast, making it unlikely that human genes in YACs will be transcribed. Even if some sequences are randomly transcribed, the resulting mRNA is unlikely to be translated and will be destroyed and recycled.

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Protocol for Extracting and Purifying YACs

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Proper Experimental Methods

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When extracting DNA from yeast for sequencing purposes, it is crucial to purify the DNA from proteins, mRNA, and linear DNA. This ensures that the native yeast DNA is separated from the circular DNA of the YAC. A detailed guide on the extraction and purification of circular YACs can be found in the paper by Noskov et al. (2010).

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Creation of YACs and Their Application

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TAR Cloning and Chemical Synthesis of Genomes

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The creation and introduction of YACs into yeast cells have been detailed in the method described by V Larionov et al. (1997). This method, known as TAR (Transformation-Associated Recombination), was pivotal in the creation of the chemically synthesized genome of Mycoplasma genitalium, as detailed in the famous paper by Gibson et al. (2008).

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Conclusion

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YACs play a significant role in the formation of chromatin within yeast cells. Their replication and condensation processes are intricately linked with the mitotic cycle, and their circular form allows for easy manipulation during genetic engineering. Understanding these processes is crucial for further advancements in chromatin biology and synthetic biology.

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