Unknown Mutation Detection via Restriction Hybridization Method Instead of Using Next Generation Sequencing Method

U. Masood
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Abstract

In biology mutation is a change in the nucleotide sequences of the DNA of an organism Mainly there are three types of mutation: point mutation, deletion and insertions. Once the mutation has been defined allele specific oligonucleotide hybridization, amplification, heteroduplex formation method referred to as a diagnostic method some advance technique like CRISPR cas9 system is using for selected mutagenesis. Using restriction method system we can detect a mutation. Let’s say you have a DNA sample with fluorescent labeled from patient and you want to make sure that gene you are interested is in healthy gene. We can design different short fragment sequences to scan through DNA or find specific gene or mutation. The sequences scan the DNA if the sequences does not find targeted gene it does not bind to it its means that no fluorescence color appears under UV-light each different short fragment sequences is label with different colors. If the different short fragments sequence does not bind to the DNA or specific gene or area this means that there will be no color appear under UV light this part or gene will be separated from the DNA by using Restriction enzyme to do a Sanger sequencing gel electrophoresis. Result of the Sanger sequencing will provide the information about sequence of unknown part or gene of the DNA this method is easier and cost economic method instead of Next generation sequencing method [1-7].
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用限制性杂交法代替下一代测序法检测未知突变
在生物学中,突变是生物体DNA核苷酸序列的变化。突变主要有三种类型:点突变、缺失和插入。一旦突变被确定,等位基因特异性寡核苷酸杂交、扩增、异双工形成方法被称为诊断方法,一些先进的技术如CRISPR cas9系统被用于选择性诱变。利用限制法系统可以检测到突变。假设你有一个病人的DNA荧光标记样本你想确保你感兴趣的基因是健康的基因。我们可以设计不同的短片段序列来扫描DNA或找到特定的基因或突变。序列扫描DNA,如果序列没有找到目标基因,则不与之结合,这意味着在紫外线照射下不会出现荧光颜色,每个不同的短片段序列被标记为不同的颜色。如果不同的短片段序列没有与DNA或特定的基因或区域结合,这意味着在紫外线下不会出现颜色,这部分或基因将通过使用限制性内切酶进行桑格测序凝胶电泳从DNA中分离出来。Sanger测序结果将提供DNA未知部分或基因的序列信息,这种方法比Next generation测序方法更简单,成本更经济[1-7]。
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