MRNA capture sequences, synthesis method of capture carrier and production method of high-throughput single-cell sequencing library

Abstract

The invention provides mRNA capture sequences. The mRNA capture sequences comprise universal primers, rare enzyme cutting sites, cell tags, random molecular tags and PolyT sequences, wherein by introducing rare enzyme cutting site sequences, sticky ends are provided for subsequent sequencing connector connection, so that two ends of a cDNA double strands are connected with two different sequencingconnectors. A synthesis method of a capture carrier used for mRNA capture and a production method of a high-throughput single-cell sequencing library are further provided. According to the provided capture carrier, by synthesizing the mRNA capture sequences of the rare enzyme cutting site sequences in situ and introducing the mRNA capture sequences of the rare enzyme cutting site sequences on a base material, the provided capture carrier is adopted for preparing the single-cell sequencing library, the single-cell capture efficiency and the labelling efficiency of oligonucleotide tags are improved, a process of constructing the library is simplified, and the two ends of the produced cDNA double strands are connected with the two different sequencing connectors, so that it is guaranteed that one dumbbell-shaped sequencing library is only assembled with a primer and DNA polymerase, and one-to-one correspondence of the dumbbell-shaped sequencing library, the primer and the DNA polymeraseis the premise of guaranteeing single-cell real-time sequencing.

Description

Technical field [0001] The invention relates to the field of biotechnology, in particular to an mRNA capture sequence, a method for synthesizing a capture carrier, and a method for preparing a high-throughput single-cell sequencing library. Background technique [0002] Tumor is one of the major diseases that seriously affect human health. There are great differences in tumor cells from genotype to phenotype (high degree of tumor heterogeneity), and this high degree of heterogeneity is related to tumor malignancy and drug resistance. Sex, recurrence and metastasis are closely related, which is one of the root causes of difficult early diagnosis of tumors, complex clinical diagnosis and treatment, drug resistance recurrence, and poor prognosis. A comprehensive analysis of tumor heterogeneity is the key to achieving precise tumor treatment. [0003] In recent years, the emerging single-cell sequencing technology has provided the possibility to analyze tumor heterogeneity and identif...

AI Technical Summary

Problems solved by technology

Because the sequences in each microwell are different, when performing high-throughput analysis, the workload of oligonucleotide sequence library and sample grafting is extremely huge. Modification work, so detection throughput is extremely limited

In addition, the existing single-cell sequencing technology usually connects the same sequencing adapter at both ends, and it is difficult to connect two different sequencing adapters at both ends of the cDNA double-stranded template.

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