Ultrahigh-flux single-cell chromatin transposase accessibility sequencing method

Abstract

The invention discloses an ultrahigh-flux single-cell chromatin transposase accessibility sequencing method. The ultrahigh-flux single-cell chromatin transposase accessibility sequencing method comprises the following steps that firstly, transposition of a chromatin transposase accessibility genome sequence in a cell nucleus is carried out by using a specific molecular tag transposase embedding compound; one molecular marker microbead and one or more cell nucleuses are located in a separated space through a microwell plate technology or a microfluidic technology, the cell nucleuses are split under the action of a splitting solution, and a specific molecular tag sequence is connected with a molecular marker sequence on the molecular marker microbead with the help of a bridging primer; a large number of sequences are obtained through PCR amplification, and a sequencing library is constructed and obtained; and then high-throughput sequencing is carried out, specific genome openness information of millions of single cells can be obtained through one-time sequencing, the single cell sequencing throughput is greatly improved, and the development of a single-cell high-flux chromatin transposase accessibility sequencing technology based on a microwell plate is actively promoted.

Description

technical field [0001] The invention relates to the technical field of single-cell sequencing, in particular to an ultra-high-throughput single-cell chromatin transposase accessibility sequencing method. Background technique [0002] In the genome, most of the chromatin is tightly wound in the nucleus and is not transcriptionally active. The chromatin state is dynamically regulated in a cell-type-specific manner, and partially dense chromatin becomes loose in a specific cell state. These loose chromatin are called open chromatin or accessible chromatin. . By detecting the opening of cell chromatin, information about cell transcription regulation can be obtained, such as where transcription factors can bind to gene promoters, which genes in cells may be efficiently transcribed, etc. Commonly used detection methods are ATAC-seq, DNase-seq, MNase-seq, FAIRE-seq and ChIP-seq, etc. These methods are based on different principles to interrupt and mark open chromatin regions. Qu...

AI Technical Summary

Problems solved by technology

The single-cell ATAC platform based on microfluidics and micropores has high sensitivity and low contamination rate, but the analysis throughput is low. A single experiment can only process about 10,000 single-cell samples, and the cost is high

sci-ATAC-seq is limited by the number of split-pool combinations, the nuclear flux of the analysis is low and the contamination rate between cells is high

On the basis of the former, sci-ATAC-seq3 uses an untagged transposase to insert the transposon fragment and then uses two consecutive rounds of split-pool connection to add cell tags, which increases the number of combinations. The efficiency of the wheel connection sacrifices part of the sensitivity

At the same time, the nucleus pre-labeling system requires multiple rounds of split-pool process, resulting in low cell recovery efficiency (about 10%)

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