High-throughput verification method of plant tissue single cell transcriptome sequencing result

Abstract

The invention discloses a high-throughput verification method of a plant tissue single cell transcriptome sequencing result, and relates to the field of gene sequencing. The method comprises the following steps: S1, degrading plant tissues by cellulose enzymatic hydrolysate to prepare a cell suspension; S2, measuring cell concentration and cell activity in the cell suspension; S3, separating different subcell populations in the cell suspension; S4, degrading the same subcell population by the cellulose enzymatic hydrolysate; S5, performing SMART-seq library building and sequencing on a cell precipitation sample; and S6, taking the cDNA obtained by library building as a template, and detecting the expression quantity of a target gene by adopting fluorescent quantitative PCR so as to carry out verification. Cell walls are degraded twice in sequence by adopting cellulose enzymatic hydrolysate so as to separate cells of different subcellular populations, fluorescent quantitative PCR detection is performed after SMART-seq database establishment, high-throughput verification of gene expression quantity is performed on single-cell and subcellular levels, the test period is short, the test cost is reduced, and the verification operation is simple, convenient, efficient and rapid.

Description

technical field [0001] The invention relates to the technical field of gene sequencing, in particular to a method for high-throughput verification of plant single-cell transcriptome sequencing results. Background technique [0002] As a revolutionary tool, single-cell RNA sequencing (scRNA-seq) technology analyzes the transcriptome at the single-cell level. The core of single-cell research is to divide cell populations and detect differences between cell populations express genes. After obtaining high-throughput single-cell transcriptome data through scRNA-seq, it is still necessary to verify the genes identified by scRNA-seq in different sub-cell populations. The use of in situ histological verification is a popular research strategy at present, and it is also a verification of the spatial distribution and positioning of valuable molecules screened out from single-cell sequencing results and the authenticity of the results. The detection of gene expression detected by scR...

AI Technical Summary

Problems solved by technology

[0005] The present invention provides a high-throughput verification method for single-cell transcriptome sequencing results of plant tissues, so as to solve the technical problems of inconvenient operation, long test period and low throughput of current transcriptome sequencing results

Images