Team:Tsinghua

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[[Image:China.gif|15px]]中文版[[Team:Tsinghua#Tsinghua University|Back]]  
[[Image:China.gif|15px]]中文版[[Team:Tsinghua#Tsinghua University|Back]]  
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2010年清华大学iGEM项目的重点是开发新的生产方法使用抗体合成生物学。其目的是'''模拟抗体的选择和生产工艺细菌'''。
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抗体是一类神奇的物质。与抗体有关的科学问题主要分为两个方向:一是在自然科学领域,研究哺乳动物体内抗体的产生机制及其作用机理,其核心问题是,免疫系统如何利用有限的基因片段产生理论上无限多种抗体,并在特定情况下大量产生适当抗体,从而应对不断变化的外界环境。二是在工程学领域,研究抗体生产技术,其核心问题是:能否发展一套迅速、便宜生产各种高特异性抗体的系统。而本质上,这两个方向却是统一的。
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传统方法生产的抗体,比如杂交瘤细胞等技术,是昂贵和费时。杂交瘤技术,对B-细胞库构建是通过实验动物,比如老鼠;使用流式细胞仪来选择一个特异性抗体,然后用杂交瘤细胞收获抗体。
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根据合成生物学的思想,清华大学iGEM10项目希望以原核生物为基础,构建一个完全模拟免疫系统抗体生成过程的人造系统,通过该系统,研究抗体形成的有关问题;而由于原核生物本身的特点,这个系统同时具有了抗体工业化生产所必需的高效、便宜的特性。实现了科学与工程的完美统一!
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在这个项目中,我们期待在大肠杆菌细胞中用一种廉价高效的方法实现所有这些过程。整个项目可分为两个模块:
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免疫系统的抗体生成过程包括两个步骤:
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-随机产生大量多样的抗体
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-对某种特定抗体进行筛选
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因此,我们的系统由两部分组成:
====模块 I: 抗体库的构建====
====模块 I: 抗体库的构建====
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我们对大肠杆菌的基因组先进行了预修饰,对其插入了重组序列与I-Sel的酶切位点。然后,我们在质粒里插入两端有重组位点的目的片段(抗体可变区V,D,J),导入大肠杆菌,诱导后利用重组序列的交叉序列即可以重组。我们具体实验验证时,会利用荧光蛋白与抗性基因的筛选。
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使用一种高效的大肠杆菌体内重组(in-vivo recombination)方法,使细菌在增殖过程中的重组来模拟免疫B细胞发育过程中复杂的重组过程,从而达到使用单一菌株简单构建抗体库的目的。
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同时,利用数学建模工具在理论上印证此系统能产生的抗体数目,可达到与免疫系统相当的数量级。
====模块 II: 特异性抗体的筛选====
====模块 II: 特异性抗体的筛选====
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实验设计主要包括利用抗原抗体结合,通过过滤方法进行筛选的“滤纸过滤筛选法”和应用抗原抗体结合后菌体间结合强度增加的性质设计的“振荡筛选法”。前者相当于利用过滤这种最常用和简便的分离方法达到了抗体分离的目的,而后者实现了用抗生素抗性这种微生物操作中最简便的筛选方法分离抗体的目的。
 
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在抗体筛选中,免疫系统所使用的方法显然要简单、高效的多。而脱离开复杂生物系统的工业生产中的筛选方法,则更易于操作。在我们的系统中,这两个筛选策略的优势得以结合。几种基于不同思路的方法被开发出来,以实现这些目的。目前主要包括:
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1、 应用原核生物跨膜信号系统模拟免疫系统膜联抗体与抗原结合后所启动的应答机制,从而将抗体筛选转化成下游简单的营养型筛选。
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2、应用原核生物胞外呈递系统将抗体筛选转化成简单的“过滤”筛选。
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3、应用原核生物结合系统将抗体筛选转化成抗性筛选。
<html>See Details @ <a href="https://2010.igem.org/Team:Tsinghua/project" style="text-decoration: none">Project</a></html>
<html>See Details @ <a href="https://2010.igem.org/Team:Tsinghua/project" style="text-decoration: none">Project</a></html>

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Our Project

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Tsinghua iGEM 2010 Project is focused on developing a new Antibody Production Method using Synthetic Biology. The Aim is to Simulate Antibody Selection and Production Technology with Bacteria.

Traditional method of antibody production such as Hybridoma Technology is expensive and time consuming. In Hybridoma Technology, generation of B-Cell library is accomplished inside experimental animal, mouse for example; using Flow Cytometry to select a specific kind of antibody; then hybridoma would be prepared to harvest antibody.

In this project, we expect to achieve all these processes cheap and efficiently in E.coli Cell. The whole project can be divided into two modules:


MODULE I: Generation of Antibody Library

We pre-modified the genome of E. coli by inserting a sequence with the I-Sel restriction sites. Then, into the plasmid of E. coli we inserted in both ends with two fragments which contain the recombinant sites (antibody variable region V, D, J). After being induced, recombination can be achieved by the crossed sequence. Specifically, as to we experiment, we will use fluorescent protein and resistance genes to testify.

MODULE II: Selection of Specific Antibody

Experimental design includes use of antigen-antibody binding, filtering through the filter called the "filtering and screening" and the application of the property as increased strength of cell adhesion caused by antigen-antibody binding to design the method called the "oscillation screening". The former is equivalent to using filters which is the most common and simple way of separating to achieve the purpose of separate and screen antibodies, while the latter one achieves the using of antibiotic resistance, which is the simplest and most convenient method in microbial operations, to separate and screen antibodies.

See Details @ Project

我们的项目

China.gif中文版Back

抗体是一类神奇的物质。与抗体有关的科学问题主要分为两个方向:一是在自然科学领域,研究哺乳动物体内抗体的产生机制及其作用机理,其核心问题是,免疫系统如何利用有限的基因片段产生理论上无限多种抗体,并在特定情况下大量产生适当抗体,从而应对不断变化的外界环境。二是在工程学领域,研究抗体生产技术,其核心问题是:能否发展一套迅速、便宜生产各种高特异性抗体的系统。而本质上,这两个方向却是统一的。 根据合成生物学的思想,清华大学iGEM10项目希望以原核生物为基础,构建一个完全模拟免疫系统抗体生成过程的人造系统,通过该系统,研究抗体形成的有关问题;而由于原核生物本身的特点,这个系统同时具有了抗体工业化生产所必需的高效、便宜的特性。实现了科学与工程的完美统一!

免疫系统的抗体生成过程包括两个步骤: -随机产生大量多样的抗体 -对某种特定抗体进行筛选

因此,我们的系统由两部分组成:

模块 I: 抗体库的构建

使用一种高效的大肠杆菌体内重组(in-vivo recombination)方法,使细菌在增殖过程中的重组来模拟免疫B细胞发育过程中复杂的重组过程,从而达到使用单一菌株简单构建抗体库的目的。 同时,利用数学建模工具在理论上印证此系统能产生的抗体数目,可达到与免疫系统相当的数量级。

模块 II: 特异性抗体的筛选

在抗体筛选中,免疫系统所使用的方法显然要简单、高效的多。而脱离开复杂生物系统的工业生产中的筛选方法,则更易于操作。在我们的系统中,这两个筛选策略的优势得以结合。几种基于不同思路的方法被开发出来,以实现这些目的。目前主要包括: 1、 应用原核生物跨膜信号系统模拟免疫系统膜联抗体与抗原结合后所启动的应答机制,从而将抗体筛选转化成下游简单的营养型筛选。 2、应用原核生物胞外呈递系统将抗体筛选转化成简单的“过滤”筛选。 3、应用原核生物结合系统将抗体筛选转化成抗性筛选。

See Details @ Project

О проекте

Russia.gifРусскийBack


Tsinghua iGEM 2010 Project is focused on developing a new Antibody Production Method using Synthetic Biology. The Aim is to Simulate Antibody Selection and Production Technology with Bacteria.

Traditional method of antibody production such as Hybridoma Technology is expensive and time consuming. In Hybridoma Technology, generation of B-Cell library is accomplished inside experimental animal, mouse for example; using Flow Cytometry to select a specific kind of antibody; then hybridoma would be prepared to harvest antibody.

In this project, we expect to achieve all these processes cheap and efficiently in E.coli Cell. The whole project can be divided into two modules:


MODULE I: Generation of Antibody Library

MODULE II: Selection of Specific Antibody

See Details @ Project

Notre Projet

France.gifFrançaisBack

Tsinghua iGEM 2010 Project is focused on developing a new Antibody Production Method using Synthetic Biology. The Aim is to Simulate Antibody Selection and Production Technology with Bacteria.

Traditional method of antibody production such as Hybridoma Technology is expensive and time consuming. In Hybridoma Technology, generation of B-Cell library is accomplished inside experimental animal, mouse for example; using Flow Cytometry to select a specific kind of antibody; then hybridoma would be prepared to harvest antibody.

In this project, we expect to achieve all these processes cheap and efficiently in E.coli Cell. The whole project can be divided into two modules:


MODULE I: Generation of Antibody Library

MODULE II: Selection of Specific Antibody

See Details @ Project

Unser Projekt

Germany.gifDeutschBack

Tsinghua iGEM 2010 Project is focused on developing a new Antibody Production Method using Synthetic Biology. The Aim is to Simulate Antibody Selection and Production Technology with Bacteria.

Traditional method of antibody production such as Hybridoma Technology is expensive and time consuming. In Hybridoma Technology, generation of B-Cell library is accomplished inside experimental animal, mouse for example; using Flow Cytometry to select a specific kind of antibody; then hybridoma would be prepared to harvest antibody.

In this project, we expect to achieve all these processes cheap and efficiently in E.coli Cell. The whole project can be divided into two modules:


MODULE I: Generation of Antibody Library

MODULE II: Selection of Specific Antibody

See Details @ Project

Nuestro Proyecto

Spain.gifEspañol Back

Tsinghua iGEM 2010 Project is focused on developing a new Antibody Production Method using Synthetic Biology. The Aim is to Simulate Antibody Selection and Production Technology with Bacteria.

Traditional method of antibody production such as Hybridoma Technology is expensive and time consuming. In Hybridoma Technology, generation of B-Cell library is accomplished inside experimental animal, mouse for example; using Flow Cytometry to select a specific kind of antibody; then hybridoma would be prepared to harvest antibody.

In this project, we expect to achieve all these processes cheap and efficiently in E.coli Cell. The whole project can be divided into two modules:


MODULE I: Generation of Antibody Library

MODULE II: Selection of Specific Antibody

私たちのプロジェクト

Japan.gif日本語 Back

清華せいさつ2010プロジェクトは、新しい抗体の生産方法は、合成生物学を使用して開発に焦点を当てています。 目的には をシミュレート抗体の選択と生産技術細菌です

ハイブリドーマ技術などの抗体産生の伝統的な方法は、コストと時間がかかります。ハイブリドーマ技術では、B細胞ライブラリの生成は実験動物内で、たとえばマウスが、抗体の特定の種類を選択するフローサイトメトリーを使用して、次にハイブリドーマは、収穫抗体を準備される実行されます。

このプロジェクトでは、我々は安く、大腸菌細胞を効率的に処理するすべてのこれらを実現する予定です。プロジェクト全体は、2つのモジュールに分割することができます:

モジュール I: 建設ののライブラリ

モジュール II: フィルタの特定の抗体