基因克隆和PCR之间的差异

关键区别 - 基因克隆与PCR

The synthesis of many copies of脱氧核糖核酸从特定的DNA片段中称为DNA扩增。有两个主要的DNA扩增过程，即基因克隆和PCR。基因克隆和PCR之间的关键区别是，基因克隆produces the multiple copies of a specific gene体内by constructing a recombinant DNA and growing inside a host bacterium while PCR produces millions of copies of a specific DNA fragment体外undergoing repeated cycles of denaturation and synthesis.

内容
1.Overview and Key Difference
2.What is Gene Cloning
3.What is PCR
4。并排比较 - 基因克隆与PCR
5。概括

What is Gene Cloning?

基因克隆是一种用于定位和繁殖特定基因的技术genomic DNAof an organism through the construction of重组DNA。Genomic DNA contains thousands of differentgenes编码为蛋白质。提取DNA时，它包含所有可能的基因。基因克隆技术使从总DNA中检测了特定基因。因此，基因克隆是分子生物学的重要工具。

Making of a基因组库of an organism is essential in gene cloning if there is no clue about the location of the relevant gene in the DNA. A genomic library is made using the following steps.

步骤1：Extraction of the total DNA from an organism which contains the desired gene.

第2步：提取的DNA的限制消化以产生较小的可管理片段。限制性核酸内切酶促进了这一步骤。

Step 3:Selection of a suitable vector and opening the向量DNA使用相同的限制性核酸内切酶。细菌质粒通常用作携带异物DNA的载体。质粒are small circles of DNA located within bacteria.

Step 4:将载体DNA和碎片DNA的结合以产生重组DNA分子。此步骤由DNA连接酶控制。

Step 5:Transferring of recombinant DNA molecules into host bacteria. This step is known as transformation, and is done using a heat shock.

Step 5:Screening of transformed bacterial cells on a culture medium. A mixed population of transformed and nontransformed host cells is obtained at the end of the transformation process. As gene of interest includes only in transformed host cells. Hence, it is necessary to select transformed cells. The selection is made using selective media which contain antibiotics. Only the transformed cells grow on this screening medium enabling the selection.

Step 6:Growing of bacteria to produce a gene library. In this step, the transformed host cells are introduced into fresh culture media which provides optimum growth requirements. Total colonies on the culture plates represent the genomic library of that organism.

步骤7：The recombinant DNA molecule containing the gene of interest must be screened from thousands of cloned fragments of recombinant DNA. It can be accomplished by the use of probes which mark the specific gene or the specific protein results from that gene.

一旦从总菌落中鉴定出含有细菌菌落的感兴趣基因，就可以制作数百万个含有该基因的重组质粒的副本。

基因克隆用于建立基因文库，生产特殊的蛋白质，维生素，抗生素，激素，测序和映射基因组，并在取证中形成多个个体DNA副本。

什么是PCR？

聚合酶链反应(PCR) is a technique which generates a large number of copies of a particular DNA fragment. Exponential amplification of a specific DNA sequence is obtained by PCR under体外条件。该技术是分子生物学中非常强大的工具，因为它可以将少量DNA样品繁殖成可用量。PCR是由Kary Mullis在1983年引入的，这项获奖的发明在分子生物学方面取得了巨大进步。

PCR技术遵循重复的PCR反应，如图02所示。一个PCR反应由三个不同温度下发生的三个主要步骤组成。在94处DNA的双链滞留⁰C，68的底漆退火⁰C和链伸长在72⁰C. Therefore, when PCR is performed, temperature fluctuation should be highly maintained for proper replication. PCR is performed in a PCR machine inside PCR tubes. PCR tubes are loaded with correct PCR mixtures containing template DNA, Taq polymerase, primers, dNTPs and buffer. Denaturing of double stranded sample DNA into single stranded DNA is done by breaking the hydrogen bonds between complementary bases at 94 – 98⁰C. Then single strands of template DNA are exposed for primers. A pair of primers (forward and reverse) should be provided, and they should be thermostable to tolerate high temperatures. Primers are single stranded short DNA sequences complementary to ends of the target DNA fragment. Synthetic primers are used in PCR. Primers bind with the complementary bases of sample DNA and initiate the synthesis of a new strand. This step is catalyzed by an enzyme called Taq polymerase; a thermostable DNA polymerase enzyme isolated fromThermus auqaticus。当提供引物和核苷酸（构建块）时，TAQ聚合酶构建了DNA与模板DNA互补的新线。在PCR程序的末尾，使用凝胶电泳观察到扩增的DNA片段。如果需要进一步分析，则从凝胶中纯化PCR产物。

PCR对于诊断和监测遗传和获得性疾病，犯罪分子的识别（在取证领域），研究靶向DNA的结构和功能非常有用科学家中的医学和分子生物学研究实验室中的常规实验室技术，因为它具有多种应用。

What is the difference between Gene Cloning and PCR?

Gene Cloning vs PCR
基因克隆是制作特定基因多副本的过程体内通过重组DNA并转化为宿主细菌。	The PCR technique produces multiple copies of a particular DNA sequence体外通过反复的PCR反应循环。
构建重组DNA的要求
产生重组DNA以定位基因。	未产生重组DNA。
需要劳动
This process is labour intensive.	不需要密集的劳动。
In vivo or In vitro process
Construction of recombinant DNA is体外and the amplification of DNA is体内。	The amplification of DNA happens completely体外。

概括– Gene Cloning vs PCR

基因克隆和PCR是用于DNA扩增的两种方法。PCR是一个体外在不使用重组DNA和宿主生物体的情况下，可以使特定DNA片段的多个DNA的多副本的DNA副本。基因克隆主要是体内process which results in multiple copies of an interested gene inside the host organism via construction of recombinant DNA. This is the difference between Gene cloning and PCR.

Reference:
1.格里菲思,安东尼·摩根富林明。”一个特定基因克隆。” Modern Genetic Analysis. U.S. National Library of Medicine, 01 Jan. 1999. Web. 22 Feb. 2017
2.“聚合酶链反应（PCR）。”国家生物技术信息中心。美国国家医学图书馆网络。2017年2月22日

Image Courtesy:
1.“图17 01 06”由CNX OpenStax -（CC由4.0）viaCommons Wikimedia
2. “PCR” By Madprime – Own work（CC BY-SA 3.0）viaCommons Wikimedia