Method for quickly amplifying target genes from genome DNA

Abstract

The invention relates to the field of gene engineering, in particular to a method related to gene cloning. The method for quickly amplifying target genes from a genome DNA comprises the following steps of: (1) preparing the genome DNA; (2) designing amplification primers of gene exons; (3) carrying out exon amplification for respectively amplifying each exon; and (4) amplifying gene segments. In the invention, by using a special primer design method, required exons are firstly amplified from the genome and then purified and mixed, and the primers at both ends are used for amplifying the required target genes in a Touch-down program. The gene splicing method avoids the preparation of RNA and a process of synthesizing cDNA by reverse transcription, thus the gene cloning can be quickly and accurately finished within several hours. Meanwhile, any cDNA sequence the sequence information of which is known can be cloned by applying the method, and the amplification length is not limited.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a method for gene cloning. Background technique [0002] Currently, there are two commonly used methods for amplifying target genes: PCR amplification and gene synthesis. Traditional PCR amplification is cumbersome, because the genome contains introns, and the low copy number of some special genes, plus some other uncertain factors, it is almost impossible to directly amplify the target gene from genomic DNA , so it is generally extracted from tissue RNA, reverse-transcribed to synthesize cDNA, and then the target gene is amplified by traditional PCR technology. However, the RNA extraction process has strict requirements and is easy to fail; in addition, some tissues containing the target gene are difficult to obtain, so this method has certain limitations. Although the target gene can be obtained by chemical synthesis, the cost of synthesis will be very expensive if the requ...

AI Technical Summary

Problems solved by technology

Traditional PCR amplification is cumbersome, because the genome contains introns, and the low copy number of some special genes, plus some other uncertain factors, it is almost impossible to directly amplify the target gene from genomic DNA , so it is generally extracted from tissue RNA, reverse-transcribed to synthesize cDNA, and then the target gene is amplified by traditional PCR technology

However, the RNA extraction process has strict requirements and is easy to fail; in addition, some tissues containing the target gene are difficult to obtain, so this method has certain limitations

Although the target gene can be obtained by chemical synthesis, if the required gene fragment is long, the cost of synthesis will be very expensive

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