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Towards construction of synthetic ri...

Li, Jun.

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Towards construction of synthetic ribosomes and a self-replicating system.

紀錄類型:	書目-電子資源 : Monograph/item
正題名/作者:	Towards construction of synthetic ribosomes and a self-replicating system./
作者:	Li, Jun.
出版者:	Ann Arbor : ProQuest Dissertations & Theses, : 2014,
面頁冊數:	104 p.
附註:	Source: Dissertation Abstracts International, Volume: 75-10(E), Section: B.
Contained By:	Dissertation Abstracts International75-10B(E).
標題:	Biochemistry. -
電子資源:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3626824
ISBN:	9781321018769

Towards construction of synthetic ribosomes and a self-replicating system.
Li, Jun.

Towards construction of synthetic ribosomes and a self-replicating system. - Ann Arbor : ProQuest Dissertations & Theses, 2014 - 104 p.

Source: Dissertation Abstracts International, Volume: 75-10(E), Section: B.

Thesis (Ph.D.)--Harvard University, 2014.

This item is not available from ProQuest Dissertations & Theses.

In 2006, the Church Group, using biochemical approaches, hypothesized that &sim;151 biomolecular components from Escherichia coli and its bacteriophages may be sufficient to enable rapid and accurate self-replication in vitro. However, efforts to construct such a system are precluded by our inability to sufficiently co-regenerate these 151 biomolecular components (or the components of ribosome---the key player of protein translation) and the inability to assemble E. coli ribosomes under conditions that are compatible with in vitro transcription and translation and also the lacking of evidence that functionally active ribosomes can be reconstituted from in vitro synthesized proteins and RNAs.

ISBN: 9781321018769Subjects--Topical Terms:

518028
Biochemistry.

Towards construction of synthetic ribosomes and a self-replicating system.
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500 $a Source: Dissertation Abstracts International, Volume: 75-10(E), Section: B.
500 $a Adviser: George M. Church.
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520 $a In 2006, the Church Group, using biochemical approaches, hypothesized that &sim;151 biomolecular components from Escherichia coli and its bacteriophages may be sufficient to enable rapid and accurate self-replication in vitro. However, efforts to construct such a system are precluded by our inability to sufficiently co-regenerate these 151 biomolecular components (or the components of ribosome---the key player of protein translation) and the inability to assemble E. coli ribosomes under conditions that are compatible with in vitro transcription and translation and also the lacking of evidence that functionally active ribosomes can be reconstituted from in vitro synthesized proteins and RNAs.
520 $a In this study, I addressed these problems in three aspects. First, I selected a well-defined cell-free system--- Protein synthesis Using Recombinant Elements (PURE) system, as the platform to construct the self-replicating system and improved the PURE system productivity in the purpose of generating sufficient amount of protein for self-replicating. I added or adjusted the concentration of a variety of factors that affect transcription and translation and achieved a boost in PURE system productivity of more than 5 fold. The work provides a more efficient defined in vitro transcription and translation system and a deeper understanding of system efficiency limitation factors. Second, I demonstrated that the 54 E. coli ribosomal proteins can be individually expressed and co-regenerated in it. Third, in contrast to conventional non-physiological methods, I developed new methods for constructing E. coli ribosomes at physiological conditions by integrating ribosomal RNA modification, cofactor facilitated ribosome assembly and in vitro transcription and translation into a one-step incubation procedure. I also showed that fully synthetic functionally active 30S ribosome can be constructed from in vitro transcribed 16S rRNA and in vitro translated 30S ribosomal proteins. Our integrated strategy solved critical barriers to construct synthetic ribosomes and synthetic life and also pave the road to build a protein based self-replicating system.
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