Massively Parallel Signature Sequencing MPSS

1
Massively Parallel Signature Sequencing (MPSS)
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Attachment of cDNA to microbeads via unique tags
AAAAAAAAA
mRNA from biological samples
x
AAAAAAAAA TTTTTTTTTT
cDNA
Unique tags (16.7M)
(20 base)
Unique complementary Anti-tag
PCR
attached
Result cDNA derived from each mRNA molecule
a single microbead
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(No Transcript)
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Micro-Bead Monolayer
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Parallel signature sequencing
NNNN
NNNN
4321
NNNN
Determine the overhang -NNNN with sequencing
cycles
NNNN
NNNN
8765
NNNN
Signature NNNNNNNN, known sequences of 20 bases
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Determine the overhang -NNNN with sequencing
cycles
e.g. N at position 1 Fluorescence-labeled
decoder for nucleotide Fluorescence-labeled
decoder for nucleotide Fluorescence-labeled
decoder for nucleotide Fluorescence-labeled
decoder for nucleotide
G
A
T
C
Call for base at position 1
G
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Track the positions of microbeads and capture
the fluorescence intensity for each decoder (G,
A, T,C) at each base position
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NNNN
NNNN
4321
NNNN
NNNN
NNNN
4321
NNNN
NNNN
NNNN
4321
NNNN
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(75,179)
G
(188,358)
G
1
(75,179)
(188,358)
10
(75,179)
G
A
(188,358)
G
A
1
2
(75,179)
(188,358)
11
(75,179)
G
A
T
(188,358)
G
A
T
1
2
3
(75,179)
(188,358)
12
(75,179)
G
A
T
C
(188,358)
1
2
3
4
(75,179)
(188,358)
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Stepping in Four Bases
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(75,179)
G
A
T
C
T
(188,358)
G
1
2
3
4
5
(75,179)
(188,358)
15
HUMFTE1A
G
A
T
C
T
T
A
C
C
C
G
T
G
A
C
A
A
A
T
A
HSRNPA1
G
T
G
A
C
G
C
T
G
A
A
T
A
A
T
A
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
(75,179)
(188,358)
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Sequencing a Specific Bead Address
G A T C C C A G T G A A T C C
T G G G T
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(No Transcript)
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(No Transcript)