used. The first development of single-cell Hi-C used manual sepa-
ration and labeling of amplified ligation products from single nuclei
to recover 10,000–30,000 ligation events per cell [70].1.5 Multiomics of
Single Cell
Single-cell genome, epigenome, and transcriptome sequencing
methods have been separately developed. However, the measure-
ments and analyses of just one molecular type from individual cells
provide only incomplete information because a cellular state is
determined by the complex interplay of multiple molecules within
its genome, epigenome, transcriptome, and proteome. To more
comprehensively and accurately analyze the mechanism by which
these molecules regulate each other, these omic methods need to
be performed in the same single cell, named multiomics. A few of
such multiomics approaches [8, 71–74] have been proposed and
enable the generation of mechanistic models relating (epi)genomic
variation and transcript/protein expression dynamics, which in
turn allow a more detailed exploration of cellular behavior in health
and disease.
To date, three types of strategies have been developed that
permit different combinations of genome, transcriptome, and
methylome sequencing for single cells. The first two approaches
allow parallel interrogation of genomes and transcriptomes for
single cells, including gDNA-mRNA sequencing (DR-seq) [71]
and genome and transcriptome sequencing (G&T-seq)
[72, 73]. For DR-seq, genomic DNA (gDNA) and mRNA present
in a single cell’s lysate are preamplified simultaneously before
splitting the reaction in two for parallel gDNA (using a modified
multiple annealing and looping-based amplification cycles (MAL-
BAC) approach [75]) and mRNA library preparation (using a
modified CEL-seq approach [35]) and subsequent sequencing.
For G&T-seq, mRNA is physically separated from gDNA using
oligo-dT-coated beads to capture and isolate the polyadenylated
mRNA molecules from a fully lysed single cell. The mRNA is then
amplified using a modified Smart-seq2 protocol [37, 76], while
gDNA can be amplified and sequenced by a variety of methods. The
second strategy enables the measures of both DNA methylation
and transcript abundance for the same single cells sampled from
dynamic and heterogeneous cell populations. The first such
method, single-cell methylome and transcriptome sequencing
(scMT-seq) [74], is an extension to the G&T-seq protocol in
which mRNA is captured, amplified, and sequenced as before.
However, the isolated gDNA of the single cell undergoes bisulfite
sequencing, rather than whole genome amplification, allowing par-
allel analysis of genome-wide DNA methylation and transcriptome-
wide gene expression from the same single cell. The third strategy
extended the second one to simultaneously probe the copy number
variation, DNA methylation, and transcript levels of the same single
cell, which are implemented in scTrio-seq [8]. The scTrio-seq334 Yungang Xu and Xiaobo Zhou