Novel methylation library-establishing kit and application thereof

Abstract

The invention relates to the technical field of molecular biology and particularly relates to a novel methylation library-establishing kit and an application thereof. The novel methylation library-establishing kit at least comprises a bisulfate conversion reagent, a sticky end joint, ligase, a DNA extending reagent and a DNA amplifying reagent. The novel methylation library-establishing kit can beapplied to a sample with a low initial mass (as low as 6pg) and suitable for genome DNA with relatively poor mass extracted from FFPE samples and the like, so that the loss of sample information caused by DNA breakage during drug treatment is greatly reduced; and a double-chain DNA joint with a sticky end can be directly introduced by virtue of relatively economic and practical common ligase, sothat the accuracy of data can be improved, the diversity of samples can be preserved to a great extent, and the library deviation property caused by human factors can be reduced.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a novel methylation library construction kit and its application. Background technique [0002] Methylation is one of the most common epigenetic modification methods, which regulate gene expression without changing the nucleic acid sequence. DNA methylation usually refers to the addition of a methyl group to the fifth carbon atom of cytosine under the mediation of DNA methyltransferase to make it into 5-methylcytosine (5-mC ). Methylation modification plays an important role in gene imprinting, gene expression regulation, X chromosome inactivation, cancer occurrence and development, etc. [0003] Abnormal DNA methylation can participate in the formation of tumors by affecting the expression of proto-oncogenes and tumor suppressor genes. When tumors occur, tumor suppressor genes usually undergo overall hypermethylation, especially in the CpG island region, thereby inh...

AI Technical Summary

Problems solved by technology

[0020] The method of self-ligation into a circle requires the use of circligase. First, the enzyme will preferentially perform intramolecular circularization, so if it is necessary to access adapters for pre-amplification, the relative linking efficiency will be greatly reduced.

Second, the enzyme is sequence-selective for the ligation of target molecules. It has no ligation ability for molecules with C at the 3' end, and the ligation efficiency for molecules with A\T\C at the 5' end is much lower than that of 5' Molecules that end in a G, so there is a severe bias

Third, the enzyme is expensive

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