SALP, a new single-stranded DNA library preparation method especially useful for the high-throughput characterization of chromatin openness states

Wu, Jian; Dai, Wei; Wu, Ling; Wang, Jinke

doi:10.1101/206854

Cited by 6 publications

(13 citation statements)

References 37 publications

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“…To address these issues, we have recently developed a new ssDNA‐based NGS library construction technique, named Single strand Adaptor Library Preparation (SALP) . This technique simplified the procedure of ssDNA‐based NGS library construction and reduced the cost.…”

Section: Introductionmentioning

confidence: 99%

“…This technique simplified the procedure of ssDNA‐based NGS library construction and reduced the cost. In addition, this technique allows the high‐efficiency, high‐throughput and low‐bias NGS library construction by using special barcoded Tn5 adaptors . Because the SALP is very suitable for the NGS library construction of highly degraded DNAs, we conceived that this technique should be very applicable to construct cfDNA NGS library, which may provide a new useful tool for the analysis of cfDNA.…”

Section: Introductionmentioning

confidence: 99%

“…To address these issues, we have recently developed a new ssDNA-based NGS library construction technique, named Single strand Adaptor Library Preparation (SALP). 19 This technique simplified the procedure of ssDNA-based NGS library construction and reduced the cost. In addition, this technique allows the high-efficiency, high-throughput and low-bias NGS library construction by using special barcoded Tn5 adaptors.…”

Section: Introductionmentioning

confidence: 99%

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Decoding genetic and epigenetic information embedded in cell free DNA with adapted SALP‐seq

Dai

et al. 2019

Intl Journal of Cancer

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Cell free DNA (cfDNA) in human plasma carries abundant information, which has therefore been the key sample for noninvasive prenatal testing (NIPT) and liquid biopsy. Especially by using the rapidly developed next‐generation sequencing (NGS) techniques, the genetic and epigenetic information embedded in cfDNA has been effectively and extensively decoded. In this process, a key step is to construct the NGS library. Due to its high degradation, the single strand‐based method was reported to be more qualified to construct the NGS library of cfDNA. In order to develop a new simple method for this application, this study adapted our recently developed single strand adaptor library preparation (SALP) method for constructing an NGS library of cfDNA. In the improved method, cfDNA was firstly denatured into single strands and then ligated with a single strand adaptor (SSA) that had a 3′ overhang of 3 random bases by using T4 DNA ligase. The SSA‐ligated DNA was converted into double‐stranded DNA with an additional adenine at the other end by polymerizing with Taq polymerase. Next, a barcode T adaptor (BTA) was ligated to this end. Finally, the cfDNA ligated with two adaptors was amplified with the Illumina‐compatible primers for NGS. Using the method, this study successfully sequenced 20 cfDNA samples from 16 esophageal cancer patients and 4 healthy people. By bioinformatics analysis, this study found the genetic and epigenetic difference between patients and healthy people and identified 23 epigenetic and 28 genetic altered esophageal cancer‐specific genes.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Decoding genetic and epigenetic information embedded in cell free DNA with adapted SALP‐seq

Dai

et al. 2019

Intl Journal of Cancer

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“…When applied to cfDNA, CATE-seq is beneficial to identify disease-causing mutations in liquid biopsy. In this study, the NGS library construction process of CATE-seq was designed to preferentially use our recently developed SALP method (36). This is a single-stranded library construction method, which is qualified to construct NGS library of any sources of DNA samples, especially high-degraded DNA such as cfDNA.…”

Section: Discussionmentioning

confidence: 99%

“…The NGS library was constructed using a SALP method developed by our lab (36). A single-strand adaptor (SSA) was prepared by annealing two oligonucleotides, SA and SA-3N (Table S5).…”

Section: Dna Sequencingmentioning

confidence: 99%

CRISPR-assisted targeted enrichment-sequencing (CATE-seq)

Xia

Zhang

et al. 2019

Preprint

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The current targeted sequencing of genome is mainly dependent on various hybridization-based methods. However, the hybridization-based methods are still limited by the intrinsic shortcomings of nucleic acid hybridization. This study developed a new CRISPR-based targeted sequencing technique, CRISPR-assisted targeted enrichment-sequencing (CATE-seq). In this technique, the input genomic DNA (gDNA) was firstly bound by a complex of dCas9 and capture sgRNA (csgRNA). The DNA-dCas9-csgRNA complex was then captured on magnetic beads through an easy room-temperature annealing between a short universal capture sequence (24 bp) at the 3′ end of csgRNA and capture oligonucleotide coupled on magnetic beads. The enriched DNAs were finally analyzed by next generation sequencing. Using this technique, three different scales of targeted enrichments were successfully performed, including enriching 35 target exons of 6 genes from 6 gDNA samples with 54 csgRNAs, 339 target exons of 186 genes from 9 gDNA samples with 367 csgRNAs, and 2031 target exons of 451 genes from 2 gDNA samples with 2302 csgRNAs. This technique has several significant advantages over the current hybridization-based methods, including high simplicity, specificity, sensitivity, throughput, and scalability.

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Finding new cancer epigenetic and genetic biomarkers from cell-free DNA by combining SALP-seq and machine learning: esophageal cancer as an example

Liu

Xia

et al. 2020

Preprint

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Background: Cancer is an important public health problem worldwide and its early diagnosis and effective prognosis are critical for its treatment. In recent years, as a good material for cancer liquid biopsy, plasma cell-free DNA (cfDNA) has been widely analyzed by next generation sequencing (NGS) for finding new molecular markers for cancer diagnosis such as size, methylation and end coordinate. However, the current studies did not still involve esophageal cancer (ESCA), a main cancer that seriously threatens human health and life in China. Here we therefore tried to find new biomarkers for this cancer from cfDNA. Materials & methods: Thirty cfDNA samples from 26 ESCA patients and 4 healthy people were used to construct the NGS libraries and sequenced by using SALP-seq. The sequencing data were analyzed with variant bioinformatics methods for finding ESCA molecular biomarkers. Results & conclusion: We identified 103 epigenetic markers (including 54 genome-wide and 49 promoter markers) and 37 genetic markers for ESCA. These markers provide new molecular biomarkers for ESCA diagnosis, prognosis and therapy. Importantly, this study provides a new pipeline for finding new molecular markers for cancers from cfDNA by combining SALP-seq and machine learning. Finally, by finding new molecular markers for ESCA from cfDNA, this study sheds important new insights on the clinical worth of cfDNA. 2 the incidence of cancer has not changed significantly before, cancer mortality has continued to decline, not only because of the development of medical standards, but also for preventive screening.Tissue biopsy is still the gold standard for diagnosing tumors, but due to the traumatic nature of patients, it brings a lot of interference to the dynamic treatment of patients. There are also many risks and ethical issues in tissue biopsy, which makes it have certain limitations. Liquid biopsy is a kind of cutting-edge technology to analyze a range of tumor material in the blood or other body fluids in a minimally invasive or noninvasive manner. The tumor material includes circulating tumor cells (CTCs), cell-free tumor DNA (ctDNA), messenger RNA (mRNA), microRNA (miRNA), and exosomes. Of these tumor biomarkers, ctDNA are most widely recognized in clinical application 4 . Comparing to ctDNA, cell-free DNA (cfDNA) is a broader term which describes DNA that is freely circulating, but is not necessarily of tumor origin. CtDNA refers to the cfDNA from tumor.CfDNA in human body fluids, such as plasma, was discovered in 1948 5 . Most of the plasma cfDNA originated from the hematopoietic system in healthy subjects, but in clinical patients (e.g. pregnancy and cancer), the related cells/tissues would release additional DNA into the plasma 6,7 . The detection of this perturbation would allow us to diagnose the abnormality for people in a noninvasive way. In recent years, methods based on the analysis of plasma cfDNA which is as an emerging technology have been largely explored for noninvasive prenatal testing (NIPT) and cancer liquid biopsy 8,9...

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SALP, a new single-stranded DNA library preparation method especially useful for the high-throughput characterization of chromatin openness states

Cited by 6 publications

References 37 publications

Decoding genetic and epigenetic information embedded in cell free DNA with adapted SALP‐seq

Decoding genetic and epigenetic information embedded in cell free DNA with adapted SALP‐seq

CRISPR-assisted targeted enrichment-sequencing (CATE-seq)

Finding new cancer epigenetic and genetic biomarkers from cell-free DNA by combining SALP-seq and machine learning: esophageal cancer as an example

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