Hierarchical DNA Memory based on Nested PCR

1
Hierarchical DNA Memory based on Nested PCR

• Satoshi Kashiwamura, Masahito Yamamoto, Atsushi
  Kameda, Toshikazu Shiba and Azuma Ohuchi,
• 8th International Workshop on DNA-based
  Computers, volume 2568 of LNCS. Springer-Verlag,
  pp. 112-123 (2003)
• Summarized by HaYoung Jang

2
Outline

• Nested PCR
• Hierarchical DNA Memory (NPMM)
• Design of Sequences
• Experimental results
• Concluding Remarks

3
Nested PCR
4
Structure of DNA Memory
Nested Primer Molecular Memory (nnn NPMM)
address part
data part
A
B
C
Re
Data
A0
B0



linker sequences
An
Bn
of hierarchy 3 of sequences in each
hierarchy n
5
Nested PCR (addressing)
A
B
C
Re
Data
B
C
Re
Data
C
Re
Data
address is represented in the primer sequences
and the order of use of them
6
Hierarchical DNA Memory

• NPMM provides a high level of data security
• NPMM has a large capacity with a high reaction
  specificity
• Enlarge the address space by using a small number
  of primer sequences
• Reduce the total errors by removing (diluting)
  the error products in each step.
• Ease of extracting the target data from MPNN

7
Hierarchical DNA Memory

• M memory capacity of NPMM.
• Data length of the sequence in the data block.
• Block number of address blocks.
• Primer number of primers in each address block.
• One base is equal to 2 bits because one base
  consists of 4 elements (A,T,G,C).

of hierarchy of sequences in each hierarchy address space of required sequences
4 100 1.00108 407
5 40 1.024108 208
6 23 1.480358108 147
7 14 1.054135108 108
8
Readout Process
A, B, C, Re 15bp Data 20bp
A
B
C
Re
Data
C0 C1 C2
A0 A1 A2
B0 B1 B2
Data0data26
9mix
3mix
27mix
target
A0
C2
B1
Re
Re
Re
all 27 NPMM units are synthesized resulting
pools are analyzed after each PCR
9
Design of Sequences

• GC_content
• Hamming distance
• 3end_complementary

10
Design of Sequences

• Designed sequences of templates and primers
• Sequence of data primers

11
Laboratory Experiments

• Extracting target data sequence using PCR

M 100 bp ladder. M' DNA marker of length 65,
50, and 35 bp. Lane 1 B0 after. Lane 2 B1
after. Lane 3 B0C0 after. Lane 4 B0C1
after. Lane 5 B1C0 after. Lane 6 B1C1 after.
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Laboratory Experiments

• Verification method

C0
Data0
C0
Data0
C0
Data0
Data0
Data1
C0
Data0
C0
Data0
35mer
amplification
Non-amplification
13
Laboratory Experiments

• Detection of amplified sequence

M DNA marker of length 65, 50, and 35 bp. Lane
1 use data000primer. Lane 2 use
data001primer. Lane 3 use data010primer. Lane 4
use data011primer.
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Laboratory Experiments

• Amplification using concatenation primer

M DNA marker of length 65, 50, and 35 bp. Lane
1 the solution set aside from the thermal cycler
at the 17th cycle. Lane 2 19th cycle. Lane 3
21st cycle. Lane 4 23rd cycle. Lane 5 25th
cycle.
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Concluding Remarks

• Hierarchical DNA memory based on nested PCR
  (NPMM)
• DNA memory with high capacity, high data security
  and high specificity of chemical reaction
• The feasibility of NPMM through some experiments