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. 2018 Sep 20;9(1):3824.
doi: 10.1038/s41467-018-06355-2.

Robust single-cell DNA methylome profiling with snmC-seq2

Chongyuan Luo  1   2 , Angeline Rivkin  1 , Jingtian Zhou  1 , Justin P Sandoval  1 , Laurie Kurihara  3 , Jacinta Lucero  4 , Rosa Castanon  1 , Joseph R Nery  1 , António Pinto-Duarte  4 , Brian Bui  4 , Conor Fitzpatrick  5 , Carolyn O'Connor  5 , Seth Ruga  6 , Marc E Van Eden  6 , David A Davis  7 , Deborah C Mash  7 , M Margarita Behrens  4 , Joseph R Ecker  8   9
Affiliations

Robust single-cell DNA methylome profiling with snmC-seq2

Chongyuan Luo et al. Nat Commun. .

Abstract

Single-cell DNA methylome profiling has enabled the study of epigenomic heterogeneity in complex tissues and during cellular reprogramming. However, broader applications of the method have been impeded by the modest quality of sequencing libraries. Here we report snmC-seq2, which provides improved read mapping, reduced artifactual reads, enhanced throughput, as well as increased library complexity and coverage uniformity compared to snmC-seq. snmC-seq2 is an efficient strategy suited for large-scale single-cell epigenomic studies.

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Conflict of interest statement

L.K. is an inventor on a patent application (US 14/384,113) submitted by Swift Biosciences Inc. that covers Adaptase. The remaining authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Improvement of single-cell methylome read mapping. a Schematics of snmC-seq workflow. Steps modified in snmC-seq2 are highlighted. RP-N and RP-H indicate random primers used in snmC-seq and snmC-seq2, respectively. Exo1 exonuclease 1, SAP shrimp alkaline phosphatase, SPRI solid-phase reversible immobilization. b Reverse read (R2) mapping is improved by reducing nucleotide concentration in random-primed DNA synthesis reaction or treatment with shrimp alkaline phosphatase, whereas forward read (R1) mapping is not affected. Each condition contained four biological replicates. c snmC-seq2 produces reverse reads with comparable base composition as MethylC-seq reads. The elements of all box-plots are defined as following—center line, median; box limits, first and third quartiles; whiskers, 1.5× interquartile range
Fig. 2
Fig. 2
Reduction of adapter dimer and short insert reads by modulating the degenerate 3′ arm of random primers. a RP-H and RP-H oligos are used for snmC-seq and snmC-seq2, respectively. b Adapter dimer and short insert reads were significantly reduced in snmC-seq2 libraries. The elements of all box-plots are defined as following—center line, median; box limits, first and third quartiles; whiskers, 1.5× interquartile range
Fig. 3
Fig. 3
snmC-seq2, snmC-seq, and MethylC-seq generate consistent methylome profiles. a Comparison of CG, CH methylation, and read coverage in the region surrounding Neurog2. L2/3 indicates combined methylome profiles of layer 2/3 excitatory neuron clusters. b Correlation of CH and CG methylation by snmC-seq2 and snmC-seq were compared for 1 kb genomic bins and CG-DMR regions, respectively. mCG, CG methylation; mCH, CH methylation
See this image and copyright information in PMC

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