The Illumina Bio-Rad

Single-Cell Sequencing Solution

Robust and scalable single-cell sequencing

For Research Use Only. Not for use in diagnostic procedures.

Gain new insight into
single-cell gene expression
With the ddSEQ Single-Cell Isolator and the SureCell WTA 3' Library Prep Kit

Single-cell RNA sequencing (RNA-Seq) provides deep This system offers:

 n end-to-end single-cell
A
insight into cell function, disease progression, and therapeutic

sequencing solution
efficacy by allowing researchers to differentiate individual cells  ensitive and unbiased
S
in heterogeneous populations. The Illumina Bio-Rad Single-Cell characterization of transcriptional
signatures
Sequencing Solution is a robust, scalable, and user-friendly
 calable cell processing to
S
workflow, developed by the industry leaders in sequencing

accommodate a wide range of

and Droplet Digital technologies, that allows transcriptome experimental designs
profiling of hundreds to tens of thousands of single cells  imple yet powerful data
S
analysis options
in a single experiment.  ccessible platform to enable
A
groundbreaking research

A streamlined and fully supported end-to-end single-cell sequencing solution

The SureCell WTA 3' Library Prep Kit provides all reagents required for single-cell isolation and barcoding using the ddSEQ Single-Cell
Isolator and library preparation with Nextera technology, while the BaseSpace Sequence Hub enables simplified run setup and push-button
bioinformatics data analysis.

< 5
min Use the ddSEQ Single-Cell Isolator to isolate tens of thousands of cells in a single day.

Scalable, flexible single-cell isolation

Process up to four samples in <5 min

Encapsulate hundreds to thousands of cells per sample and tens of thousands of cells per day
Generate stable and uniform droplets for robust cell lysis and efficient barcoding with Droplet Digital technology
 High-quality data
A A A
5x104 7,000
Average Cell Cells: 1,384
Cell Line Species Cell Type Diameter, m Doublets: 5.8% 6,000
4x104

Number of UMIs aligned

Purity: 99.1%

Mouse genes detected

HEK 293 Human Embryonic 20 5,000

to mm10 genes
kidney 3x104 4,000
NIH 3T3 Mouse Embryonic 15
3,000
fibroblast 2x104
2,000
A20 Mouse B lymphocyte 10 1x104
1,000

BJ Human Foreskin 25 0 0
fibroblast 0 1x104 2x104 3x104 4x104 5x104 0 50 100 150 200 250
Number of unique transcripts Reads per cell, x1,000
aligned to hg19 genes
B B B
8,000 1.0 10 6
R2 = 0.98
7,000 No. genes = 10,911
Cumulative fraction of genic

0.8
Median genes detected

10 5

Gene count Rep 2

6,000

5,000
UMI count

0.6 104
4,000
0.4 103
3,000

2,000 102
0.2
1,000
0.0 101
0
HEK 293 BJ NIH 3T3 A20 0 2,000 4,000 6,000 8,000 101 102 103 104 105 10 6
Cell line Cells in descending order by unique Gene count Rep 1
transcripts aligned to genes count

Unbiased, cell size agnostic RNA-Seq
HEK 293, NIH 3T3, A20, and BJ cells were processed
using the SureCell 3' WTA Library Prep Kit. A, cell sizes as
measured by the TC20 Automated Cell Counter; B, the
median number of genes detected per cell is shown.
Confident single-cell transcript identification Highly sensitive and reproducible results
HEK 293 and NIH 3T3 cells were analyzed in a mixed
species experiment. A, plot of human and mouse unique
transcripts detected in a mixed species sample. The low
percentage of cell barcodes (5.8%) containing transcripts
mapping to both human and mouse demonstrates efficient
single-cell isolation with low doublets and high purity (99%).
B, plot of cumulated fraction of genic transcripts assigned
to cell barcodes. The inflection point (knee) is used to
determine the number of barcoded cells detected, indicating
high fraction of transcripts assigned to single cells.
Replicate samples were processed and sequenced on a
NextSeq 550 run. A, sequencing reads were sub-sampled
to various reads per cell ranging from 25,000 to 200,000
reads. The median number of genes detected per cell is
plotted for each sequencing depth. B, total gene counts
for each human gene were summed across all NIH 3T3
cells. Linear regression fit of summed gene counts
between replicates from a ddSEQ M Cartridge shows high
reproducibility. Rep 1 ( ); Rep 2 ( ).

Second strand synthesis Store data and monitor

runs in real time.
TTTT Bar Read 1
Code
Automatic data Simplified
Direct cDNA Nextera tagmentation upload to BaseSpace data analysis with
Sequence Hub BaseSpace Apps

P7 Index
3 enrichment and sample indexing

P5

Sequencing-ready fragment

P7 Index Read 2 TTTT Bar Read 1 P5

Code

Rapid and streamlined library preparation Completely integrated sequencing and data analysis

 repare libraries without shearing or pre-amplification using

P  xperience streamlined, push-button data analysis with
E
Nextera technology the single-cell analysis BaseSpace app
Sequence up to 24 samples with Illumina indices Analyze, visualize, and share data easily
Detect more genes with sensitive assay chemistry Trust in secure and scalable cloud-based data analysis and storage


B
t-SNE axis 2

M/G1
M/G1
M
G2/M
G2/M
S
G1/S
G1/S

M/G1
MM
G2/M
SS
G1/S
H

30
20
10
0
10
20
30
ctattatgagacgtcggc
ctattatgagacgtcggc
ctattaatacttagatgt
ctattaatacttagatgt
aacgtgaacgtgcgagca
aacgtgaacgtgcgagca
agcacgtaccgacgcata
agcacgtaccgacgcata

30
atgaagcaaccggcgaat
atgaagcaaccggcgaat
gaaataaaagaaacaagg
gaaataaaagaaacaagg

Low
Low
gcttgtaaagaaccacgc
gcttgtaaagaaccacgc
gtcggccgcatacagact
gtcggccgcatacagact
ggtgctcgcatacggtcc
ggtgctcgcatacggtcc
cactgtatactttggcag
cactgtatactttggcag
ctcaattaatagacccaa
ctcaattaatagacccaa
caggagcagacttgtgta
caggagcagacttgtgta
aagccagccagatgtgta
aagccagccagatgtgta
cctctaacccaagccgtt
cctctaacccaagccgtt

sub-population.
aagtattcgccttgtgta
aagtattcgccttgtgta
aagtatggattgggacga
aagtatggattgggacga

20
caccacctaggtactgca
caccacctaggtactgca
gcgcgggcttgtttggat
gcgcgggcttgtttggat
gctgaggcgaatcgttat
gctgaggcgaatcgttat
cttacgtccaagaagtat
cttacgtccaagaagtat
cggtccttcttggtcggc
cggtccttcttggtcggc
gagtgatctagcaattgg
gagtgatctagcaattgg
ccgatggctgagtttggg
ccgatggctgagtttggg
accttcctgtggccgatg
accttcctgtggccgatg
ggccatacaaggttaaga
ggccatacaaggttaaga
aggttatgcggtgccgtt
aggttatgcggtgccgtt
caccacgaaggggtttca
caccacgaaggggtttca
caagtcttggatggccat
caagtcttggatggccat

Low Average
Average
cagactcagacttcagtg
cagactcagacttcagtg

10
atatacttcgcagactcg
atatacttcgcagactcg

ExpressionExpression
Expression
cactgtgaaggggtggtg
cactgtgaaggggtggtg
atagcgtcagtgcagact
atagcgtcagtgcagact
ctaggtaggttacgagca
ctaggtaggttacgagca
gagcttcgagcaaagcca
gagcttcgagcaaagcca

t-SNE axis 1

Bulletin 6855 Ver A 16-0903 1016

gccagagactcgccgtaa
gccagagactcgccgtaa
gagtgacttacgatactt
gagtgacttacgatactt
aacagcccacgcgactcg
aacagcccacgcgactcg
aacagcgactcgactgca
aacagcgactcgactgca
acccaaatccggcataga
acccaaatccggcataga

0
cttgaacatagatatttc
cttgaacatagatatttc
gcgaatatagcgcactgt
gcgaatatagcgcactgt
gatcaaacccaagaggcc
gatcaaacccaagaggcc
cggtccttggatgctgag
cggtccttggatgctgag
cgagcaatatacccacgc
cgagcaatatacccacgc

Average High
High
High
agaccccatagaatatac
agaccccatagaatatac
tagaggcagactgcgcgg
tagaggcagactgcgcgg
cttgaaagtctggtcggc
cttgaaagtctggtcggc
ccacgctgcggtacggac
ccacgctgcggtacggac
caccacatatactggcag
caccacatatactggcag
accttcgggatcggattg
accttcgggatcggattg
cggtcccatagacgaaag
cggtcccatagacgaaag
gactcggatcaacataga
gactcggatcaacataga

10
cgcatacagactcgaaag
cgcatacagactcgaaag
agaccctcgggaggtgct
agaccctcgggaggtgct
Push-button data analysis setup

tatttcagtaaaagatgt
tatttcagtaaaagatgt
ctgtgggatcaaagtaaa
ctgtgggatcaaagtaaa
agcacgtctagcctgtgg
agcacgtctagcctgtgg
cttacggggatcaggtta
cttacggggatcaggtta
caccacgctgagcaccac
caccacgctgagcaccac
cttgaatgcggtcaggag
cttgaatgcggtcaggag
and differentially expressed genes
Convenient sequencing QC metrics

ctgtggcgagcaaggtta
ctgtggcgagcaaggtta

30
ctcaattgctaagggatc
ctcaattgctaagggatc
cagacttgaattcataga
cagacttgaattcataga
Exportable gene expression matrices

gagtgactaggtagatgt
gagtgactaggtagatgt

Pub No. 1076-2016-011 Current as of 29 September 2016

cgcataatactttatttc
cgcataatactttatttc
ctaggtccacgctaagct
ctaggtccacgctaagct
taagctgtggtggccaga
taagctgtggtggccaga
ggattggccagatggcag
ggattggccagatggcag
primary and secondary analysis including:

gaaataagtctgcactgt
gaaataagtctgcactgt
aagtataggttattaaga
aagtataggttattaaga
acggacttggatacggac
acggacttggatacggac
atatacagcacgcgcata
atatacagcacgcgcata
acaaggtaccgagccgtt
acaaggtaccgagccgtt
gcgcggttggatgagtga
gcgcggttggatgagtga

20
acaaggtagaggctatta
acaaggtagaggctatta
taagctacccaataatag
taagctacccaataatag

2016 Illumina, Inc. | Bio-Rad Laboratories, Inc. All rights reserved.

ctattactattaagtaaa
ctattactattaagtaaa
ctgtggcatagaaacgtg
ctgtggcatagaaacgtg
cctctaattagttaagct
cctctaattagttaagct
ggacgactaggtgttaac
ggacgactaggtgttaac
cggtcctcatcatcggga
cggtcctcatcatcggga
gtggtgggccatttggat
gtggtgggccatttggat
gttaacaggttaaagcca
gttaacaggttaaagcca
taatagcttgaaggacga
taatagcttgaaggacga
gccagatctagcgatcaa
gccagatctagcgatcaa
actgcaagcacggggatc
actgcaagcacggggatc

10
cgaaaggaggcccgagca
cgaaaggaggcccgagca
accttcaagtatgtacag
accttcaagtatgtacag
ggccataacagcctgtgg
ggccataacagcctgtgg
acccaaggccatcactgt
acccaaggccatcactgt
Easy assignment of unique transcripts to single cells

cactgtaagtattttggg
cactgtaagtattttggg

t-SNE axis 1
aacgtgcagactgtacag
aacgtgcagactgtacag
atacttagtctggggatc
atacttagtctggggatc
Simple, yet powerful, data analysis options

tatttcttaagacgcata
tatttcttaagacgcata
aggttaagtaaatgcggt
aggttaagtaaatgcggt
aattggctaggttgttcg
aattggctaggttgttcg

hg19 RPL13

0
agtaaagctcccgagtga
agtaaagctcccgagtga
acccaagctccctaagct
acccaagctccctaagct
gcgaatatactttgtgta
gcgaatatactttgtgta

For Research Use Only. Not for use in diagnostic procedures.

atacttcgttatcaggag
atacttcgttatcaggag
caagtcagtctgtgctaa
caagtcagtctgtgctaa
The single-cell analysis BaseSpace app enables streamlined

ctgtggcatagaagcacg
ctgtggcatagaagcacg
atagcgcaccaccaagtc
atagcgcaccaccaagtc
gccgttttcttgggattg
gccgttttcttgggattg

a distinct cluster in t-SNE analysis based on gene expression profile (left). Cells
cgaaagcgaaagatgaag
cgaaagcgaaagatgaag

cells. Human cells (representing 7% of the total cell population) are identified as
attagtgatcaatgtgta
attagtgatcaatgtgta
 igh dimensional analysis for identification of subpopulations

cctctaccgatgtgtgta
cctctaccgatgtgtgta

color-coded by gene expression of human Rpl13 (right) confirm the identity of the
caccacttgctcgggatc
caccacttgctcgggatc
cgagcattggatcggcgt
cgagcattggatcggcgt

10

Powerful data analysis tools. The single-cell analysis BaseSpace app provides a
aaagaatccaagcttgaa
aaagaatccaagcttgaa

profiling of proliferating HEK 293 cells using expression of markers associated with
suite of data analysis options including heat maps and t-SNE analysis. A, cell cycle
atgaagagtaaatgtgta
atgaagagtaaatgtgta
gcgaattgctaacataga
gcgaattgctaacataga

each major cell cycle phase. B, t-SNE analysis of a mixture of HEK 293 and NIH 3T3
ctgtggtgagacaacagc
ctgtggtgagacaacagc
ggccatcaccacgccgtt
ggccatcaccacgccgtt
atacttgctgagatccgg
atacttgctgagatccgg
tcagtgcttacgagtaaa
tcagtgcttacgagtaaa
caaccgccgatgggattg
caaccgccgatgggattg
caagtcacccaaccacgc
caagtcacccaaccacgc
tccaagccacgcatagcg
tccaagccacgcatagcg
ctattacggtccgcgcgg
ctattacggtccgcgcgg
cggcgtaacgtgcataga
cggcgtaacgtgcataga
ggccatgactcgagcacg
ggccatgactcgagcacg
ctgtggtcagtggcttgt
ctgtggtcagtggcttgt
gtacagtgctaaccgtaa
gtacagtgctaaccgtaa
gcgaatgagtgataagct
gcgaatgagtgataagct
aggttacgcataacccaa
aggttacgcataacccaa
aagccatatttcgcgaat
aagccatatttcgcgaat
gctgagctgtggcaggag
gctgagctgtggcaggag
atacttcttacggatcaa
atacttcttacggatcaa
ggtgctcgaaagggtgct
ggtgctcgaaagggtgct
cgcataaacgtgaattgg
cgcataaacgtgaattgg
ctgtggctaggttgagac
ctgtggctaggttgagac
gtcctagtggtgtgctaa
gtcctagtggtgtgctaa
acggaccatagatagagg
acggaccatagatagagg
tccaagggtagggccaga
tccaagggtagggccaga
gttaacccacgcaagtat
gttaacccacgcaagtat
cttgaattaagacgaaag
cttgaattaagacgaaag
taagctggtaggcggtcc
taagctggtaggcggtcc
gagtgacagactcttacg
gagtgacagactcttacg
gcgcggtgagacagatgt
gcgcggtgagacagatgt
gactcggtcctaagtaaa
gactcggtcctaagtaaa
tcatcaagtaaaagtaaa
tcatcaagtaaaagtaaa
agtctgcctctacggtcc
agtctgcctctacggtcc
cctctagaggcccttgaa
cctctagaggcccttgaa
gcgaattggcagggtagg
gcgaattggcagggtagg
and analysis.

atccggtgctaactcaat
atccggtgctaactcaat
gccagaggtaggaagcca
gccagaggtaggaagcca
cgaaaggtcggctgtgta
cgaaaggtcggctgtgta
gcgaattatttcatccgg
gcgaattatttcatccgg
gtttcaactgcaccgatg
gtttcaactgcaccgatg
cagactctgtggcaaccg
cagactctgtggcaaccg
gcttgtgaagggtgcggt
gcttgtgaagggtgcggt
aattgggactcggcgaat
aattgggactcggcgaat
atagcgaattgggcgaat
atagcgaattgggcgaat
atccggcatagacgaaag
atccggcatagacgaaag
gaaataagatgtctgtgg
gaaataagatgtctgtgg
ctaggtttcttggttaac
ctaggtttcttggttaac
gaagggaagccagtttca
gaagggaagccagtttca
ggccatccacgcagaccc
ggccatccacgcagaccc
caaccgggacgatcagtg
caaccgggacgatcagtg
cgttattcgccttaagct
cgttattcgccttaagct
cggtccctcaatactgca
cggtccctcaatactgca
gaagggcgcataggtagg
gaagggcgcataggtagg
accttcgtcctagatcaa
accttcgtcctagatcaa
gcgcggggccatctatta
gcgcggggccatctatta
ccgatgactgcagctccc
ccgatgactgcagctccc
caagtctcatcagggatc
caagtctcatcagggatc
cgaaagttggatgggatc
cgaaagttggatgggatc
aacgtgcggcgtttgctc
aacgtgcggcgtttgctc
ggacgattcgcaaagcca
ggacgattcgcaaagcca
ccgtaatcatcattcttg
ccgtaatcatcattcttg
atagcgtgttcggactcg
atagcgtgttcggactcg
agacccaagccaagatgt
agacccaagccaagatgt
by technology leaders

aagccacgagcataagct
aagccacgagcataagct
cactgtcggcgtgtccta
cactgtcggcgtgtccta
agcacgcttgaatcggga
agcacgcttgaatcggga
ccgtaaaacgtgacggac
ccgtaaaacgtgacggac
ccgtaacggtcccttacg
ccgtaacggtcccttacg
agatgtcaccaccgcata
agatgtcaccaccgcata
acggacttcgcagccaga
acggacttcgcagccaga
atacttgatcaaaagcca
atacttgatcaaaagcca
gcgaatttcgcacactgt
gcgaatttcgcacactgt
atagcgccgtaattaaga
atagcgccgtaattaaga
aacagcgatcaattcgca
aacagcgatcaattcgca
ctaggtcatagaatatac
ctaggtcatagaatatac
agtaaaagtaaagctgag
agtaaaagtaaagctgag
caccacgccagaaattgg
caccacgccagaaattgg
cttgaatggcagcataga
cttgaatggcagcataga
cggtccgcttgtacggac
cggtccgcttgtacggac
cggcgtgggatcgagtga
cggcgtgggatcgagtga
agatgtttcgcatggcag
agatgtttcgcatggcag
ggacgaacaaggtctagc
ggacgaacaaggtctagc
caaccgacggacatgaag
caaccgacggacatgaag
gctgagaagtatgatcaa
gctgagaagtatgatcaa
gactcggtcctatcatca
gactcggtcctatcatca
ccacgccaccacttaaga
ccacgccaccacttaaga
atataccatagaagcacg
atataccatagaagcacg
caagtcagacccatactt
caagtcagacccatactt
corresponding U.S. and non-U.S. patent applications.

cttacggccgtttttggg
cttacggccgtttttggg
A complete solution built and supported

without limitation, quality control and commercial services such as reporting

expressly, by implication, or by estoppel. This product is for internal research
Illuminas expertise in library preparation and sequencing and

purchaser. No right to resell this product or any of its components is conveyed

the results of purchasers activities for a fee or other form of consideration. The

Bio-Rad Laboratories, Inc., including U.S. patents 7,772,287 and 9,216,392, and
right to use the product only to perform internal research in the field of single cell

product and its use are covered by one or more patents owned by or licensed to
analysis and with Bio-Rads ddSEQ Single-Cell Isolator for the sole benefit of the
eliminating the need to seek support from multiple vendors. Full
Bio-Rads industry-leading Droplet Digital technology to provide

workflow support is available from isolation through sequencing

The Illumina Bio-Rad Single-Cell Sequencing Solution leverages

an integrated, scalable, and fully supported single-cell RNA-Seq

purposes only and is not for use in commercial applications of any kind, including,
The purchase of this product conveys to the purchaser the limited, non-transferable
workflow. Technical support can be obtained from a single source,

Visit bio-rad.com/web/singlecell for more information.