The 3The 3rdrd Research on Research on Theorem ProvingTheorem Proving
MEC Meeting MEC Meeting
Hanyang University Hanyang University Proteome Research LabProteome Research Lab
2003. 1. 16 2003. 1. 16
Park, Ji-Yoon Park, Ji-Yoon
The 1The 1stst Year Year ResearchResearch
- Linear Implementation- Linear Implementation
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 33
Theorem Proving using Resolution Theorem Proving using Resolution Refutation Refutation
NilNil
, add R as, add R as
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 44
PrimerPrimer SequenceSequence LengthLength ModificationModificationGC ContentGC Content
( % )( % )
ST¬QST¬Q5’ - AAG CAG TAG CGA CCA 5’ - AAG CAG TAG CGA CCA
ATT GAC GCA AAT TGA ATT GAC GCA AAT TGA CGT ACG TAC GCT GAA - 3’ CGT ACG TAC GCT GAA - 3’
45 mer45 mer NoneNone 46.746.7
¬RPQ¬RPQ5’- CAT ACA ATG AAC GCA 5’- CAT ACA ATG AAC GCA
GTC AAC GCA AGG CAG GTC AAC GCA AGG CAG TTC AGC GTA CGT ACG - 3’TTC AGC GTA CGT ACG - 3’
45 mer45 mer NoneNone 51.151.1
¬P¬P 5’- CTG CCT TGC GTT GAC - 3’5’- CTG CCT TGC GTT GAC - 3’ 15 mer15 mer 5’-phosphate5’-phosphate 60.060.0
RR 5’- TGC GTT CAT TGT ATG – 3’5’- TGC GTT CAT TGT ATG – 3’ 15 mer15 mer 5’-phosphate5’-phosphate 40.040.0
¬S¬S 5’ - TGG TCG CTA CTT –3’ 5’ - TGG TCG CTA CTT –3’ 15 mer15 mer 5’-phosphate5’-phosphate 53.353.3
¬T¬T 5’ - TCA ATT TGC GTC AAT-3’ 5’ - TCA ATT TGC GTC AAT-3’ 15 mer15 mer 5’-phosphate5’-phosphate 33.333.3
The Sequence for Linear The Sequence for Linear Implementation Implementation
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 55
587
540
504
458
434
267
234
213
192
184
124/123
104
89/80
64/57/51/
21/18/11/8
75 bp75 bp
M 1 2
The Amplification of Linear The Amplification of Linear MoleculeMolecule
Fig 1. Fig 1. The amplification of linearThe amplification of linear molecule in 3% agarose gel molecule in 3% agarose gel electrophoresis electrophoresis Lane 1: PCR product with S and ¬R Lane 1: PCR product with S and ¬R Lane 2: PCR products with ¬ S and R Lane 2: PCR products with ¬ S and R Lane M is a 25 bp molecular DNA ladder Lane M is a 25 bp molecular DNA ladder
The 2The 2ndnd Year Year ResearchResearch
- Hairpin Implementation- Hairpin Implementation
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 77
Sequence for Hairpin Sequence for Hairpin Implementation Implementation
5mer5mer
5mer5mer
6 mer6 mer
¬P¬P
PP ¬Q¬Q
Q Q
RR
P
Q
S
TP
Q
S
T
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 88
PrimerPrimer Sequence(5’ → 3’)Sequence(5’ → 3’) LengthLength ModificationModificationGC ContentGC Content
( % )( % )
PP TATTAAGACTTCTTGTAGTCT TATTAAGACTTCTTGTAGTCT 21 mer21 mer 5’-Phosphate5’-Phosphate 28.528.5
QQ TCATGTTCCT TCATGTTCCT 10 mer10 mer 5’-Phosphate5’-Phosphate 40.040.0
¬P¬P TAATAAGGAATAATAAGGAA 10 mer10 mer 5’-Phosphate5’-Phosphate 20.020.0
¬Q¬Q CATGA CATGA 5 mer5 mer 5’-phosphate5’-phosphate 40.040.0
¬R¬R TCATGTTCCTTCATGTTCCT 10 mer10 mer 5’-phosphate5’-phosphate 33.333.3
The Sequence for Hairpin The Sequence for Hairpin Implementation Implementation
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 99
The Amplification of Hairpin The Amplification of Hairpin MoleculeMolecule
Fig 2. The amplification of hairpin molecule in 3%Fig 2. The amplification of hairpin molecule in 3% agarose gel electrophoresis agarose gel electrophoresis
50 bp50 bp
25 bp25 bp
The 3The 3rdrd Year Plan Year Plan
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 1111
Design of Problem Design of Problem
6 variables, 9 6 variables, 9 clausesclauses
322232122212
312131112111
323122211211
,,
,,
,,
PPPPPP
PPPPPP
PPPPPP
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One of Proof TreesOne of Proof TreesPP1111∨P∨P1212 ~P12∨~P22
P11∨~P22 P21∨P22
P11∨ P21 ~P21∨~P31
P11∨~P31 P31∨P32
P11∨ P32 ~P12∨~P32
P11∨~P12 P11∨P12
P11
~P11∨~P21 P21∨P22
~P11∨ P22~P22∨~P32
~P11∨ ~P32P31∨P32
~P11∨ P31~P11∨~P31
~P11
nnilil
20~30
20~30
2003. 1. 16 2003. 1. 16 Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ Advanced Proteome Research Lab(APRL) http://gene.hanyang.ac.kr/ 1313
Experimental ProcedureExperimental Procedure
The Problem The Problem (6var, 9clau)(6var, 9clau)
Design of Oligo SeDesign of Oligo Sequence quence
Exonuclease III Exonuclease III (remove partial (remove partial solution) solution)
Step IStep I
Step IIStep II
Step IIIStep III
Step IVStep IV
Step VStep V
HybridizationHybridization(95°C→16°C (95°C→16°C cooling down) cooling down)
Synthesis & Synthesis & ModificationModification(5’-Phosphate)(5’-Phosphate)
LigationLigation(T4 DNA Ligas(T4 DNA Ligas
e: 16°Ce: 16°C
Step VIStep VI
Step VIIStep VII
Gel Electrophoresis Gel Electrophoresis
Step VIIIStep VIII
The Final The Final SolutionSolution
* * No Self-No Self-HomologyHomology
* No Cross-* No Cross-HomologyHomology
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Forward DirectionForward Direction DNA9 Conference DNA9 Conference
Abstract Submission(2/15)Abstract Submission(2/15) Experimental Work Experimental Work
Journal Version Journal Version Biochemical Journal Biochemical Journal Biological Computing Biological Computing
6.3736.373
7.6667.666
3.161 3.161
4.493 4.493
5.545 5.545
29.491 29.491
36.242 36.242
24.595 24.595
10.89610.896
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Expected ResultExpected Result
Fragments of linear DNA migrate thFragments of linear DNA migrate through agarose gels with a mobility rough agarose gels with a mobility that is inversely proportional to the that is inversely proportional to the loglog1010 of their molecular weight of their molecular weight
Circular forms of DNA migrate in aCircular forms of DNA migrate in agarose distinctly differently from lingarose distinctly differently from linear DNAs of the same mass.ear DNAs of the same mass.
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DiscussionDiscussion