????? miRNAs ?????? ???? ???

1
?????????? ??
???? ??????? ???-
???????? ???????

• ????? miRNAs ?????? ???? ???
• ??????? small RNA sequencing
• ?? ???? ???
• ???? ?"? ??? ??????

2
MicroRNAs(miRNAs)
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
microRNAs
Non-coding RNA
Small RNA

• ????? ?? small RNA
• ?????? ????? ???? ?????? ?? ??????? ?????
• ???????? ??????? ??????? ?? ? mRNA
• miRNA ???? ????? ?? ????? ? mRNA ?????? ?? ?????
  ??????? ?"? ??????????? ?? ??? ? 3' UTR.
• ?????? ???, ???? ? miRNA ?????? ?? ????? ????
  ???? ??????

3
???? ??????? miRNA
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
miRNA ?????? ?"? RNA polymerase II ? pri-miRNA-
????? ? pri-miRNA ????? stem loop ????? ?? 60-70
???????????, ????? pre-miRNA , ????? ?? ?????
?"? ?????????? Drosha RNase III
pre-miRNA ????? ??????? ??????????
????? ,miRNA duplex RNA ?? ????? ????? 22
????????????
??? ??????? ????????, ?miRNA ,??????? ???????
RISC ???? ?? ?????? ?? ?? ?????. ?????? ????
miRNA, , ???? ????????.
4
???? ????? ???
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ????? ???????? ?????? ???? ?????? ?? ???? ????.
• ????? ???????
• ??????
• ????????
• ?????? ?????????
• ???? ??? ????? ?????? ?????? ????? ???? ????
  ????? ??????.

5
???? ??? ?????? ??????
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ??? ???? ??????? ????? ????? ????? ????? ??
  ??????
• BRCA1
• BRCA2
• ???? ??? ?????? ??????? ?????? ?????? ??? ??????
  ????? ? DNA.
• ????? ??? ??? ???? ?????? ?????? ?? ????, ??????
  ?? ?????? ????? ????? ???.
• ???? ?????? ?? ??????? ? BRCA1 ?-BRCA2 ??????
  ?????? ???? ????? ????? ?? ?????? ????.
• ?????? ???? ?????? ????? ??-50-80 ????????
  ???? ??? , ????? ?-12 -????? ???????? ?????
  ?????????? ??????.

6
miRNAs ????? ???
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ??????? ????????, ?????? ???? ????? ?????? ??????
  ???? ???? ????? ??? ?????? ?????? ??? ????? ??
  ?????? ??? ??????. ?????? ?????? ?? miRNAs ????
  ?????? ???? ????. ???? ? miRNAs ?????? ?????
  ????? ?????? ??????? ????? ??? ????????,
  ?????????, ???????? ???????? ?? ?? ?????? ??????
  ??? ????. ???? ??? miRNAs ????? ??? ???? ?"?
  ?????? ??????? ????? miRNAs ????? ??? ????? ??
  ?????? ???????. ?????? ??? ???? ?? ???? ??????
  ????? miRNAs ????? ?????? ??????? ??????
  ????????. ???? miRNAs ???????? ????? ????? ????
  ???? ????? ????.

7
Next-generation sequencing
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ??? ????? ??? ????? ??????, ???? ???????? ?????
  ????? ??????? ???????? ?? ???? ?? ????? ??????
  ????????.
• ?????? ???? DNA ?RNA ????? ???? ??? ????
  ?????????, ?? ?????? ?? Sanger sequencing ??????
  ???? ?????? ????? ???? ?? ?? ????? ????????
  ?????, ????????? ?? ???? ????? ?? ???? ????
  ?????? ???? ?????? ??????, ??????, ????????.
  ?????? ???? ???? ????? ??? ?????? ???. ???
  ??????, ?????? Sanger ????? ?????? ???? ?????
  ???? ?? ?????? ????? ????? ?????, ????? ???
  ??????? next generation sequencing ????? ??????
  ????? ???? ???? ??? ???? ????? ?????.NGS ?????
  ?? ????? ?????? ???????? ???????? ????? ??????,
  ?????? ??????? ????? ???? ?? ???? ??????.
• ??????? ??????? ?? ????? ??? ?? ?? ????, ????
  ?????? ????? ??? ????? ??????? ????? ??????
  ???????? ??????.

8
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ?????? ?????????????? ??????? ?????? miRNAs ?????
• ???? ?? ???? ?? ?? ?????? ?????????????? ??????
  ?"? ????? miRNAs ????? ????'miRDeep ,
  miRanalyzer, miRExpress, miRTAP,mirTools.
• ???????? ?? ??????? ???? ?????? miRanalyzer ?
  miRDeep2. ???? ?????? ??? ?? ??? ??????web
  server ? stand alone. ???????? ?? ??????? ?????
  ?????? ????? ??? ?? ???? ??????? ????' ???? ?????
  ???? ??????? ??????? ?????? ????? ?????.

9
???? ??????
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ????? ?????
• ????? ???????
• ????? ???? ?????? ?????? ?????? ?? ???? ????????
  ???????? ?? ???????.
• ???? ???? ????? ?? ?????? ?????? ??????-??????
• ???? ?????? ?????? ??????.

10
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ????? ?????
• ? TCGA ??? ?????? ?????? ???? 2006. ??????? ????
  ?? ????? ?? ??????? ??????? ?????? ???? ????
  ????????. ???????? ? TCGA ?????? ???? ???? ????
  ?? ??????? 500 ?????? ???? ?? ??? ?????. ???
  ???? ????? ?????? ???? ???? ?? ?? ??? ???? .
• ?? ????? ???? TCGA ?????? ?? ???? ???????. ??????
  ??? ??? ???? ?? ??? ???? ???? ???????.

11
(No Transcript)
12
miRanalyzer
13
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ?????? ?????? ???? ?????? next generation
  sequencing ???? ????? ??????? ?? ???? ????? ??
  data ????? ???? ?????????????? ??????? ?? ???? ??
  ??? ????? ??. ???? ??? ????? ?? ????? ?
  miranalyzer ????? ??????? ??? ?????? ???????
  ???? small RNA ???????? ? deep sequencing
  experiments
• ??? ?? ???? ???? input ???? ?? ????? ????? ?? ?-
  reads (?????? ?? ????? ?NGS) ????????? ?????
  ??????? ????.
• Miranalyzer ???? ?? ????? ???
• ???? ?? ?? ? miRNA ??????? ???????? ?????
  miRBase.
• ???? miRNAs ?????.

14
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• Miranalyzer ????? ?
• Weka ???? ?? ?????????? ?????? ?????? ???????
  ???????.
• Vienna RNA package ????? ?????? ????? ??????? ??
  ? RNA .
• Bowtie ??? ???? ?????? alignment ? short reads .

15
miRBase
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ??? ??? ?????? ????? ?? ?????? ??????? ??
  microRNA. ???????
• -???? ????? ???? ????? ?? microRNAs
• -????? ?? ?? ???? ? microRNAs ???????
• - ????? ??? ?? microRNA targets.
• -???? ???? ?? ????? ?miRNA ??? ????.

16
????? ????? ?? ???? ????? ?????? miRanalyzer
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
FASTQ
Perl script
Read count(RC)
Clean RC
Aligns to DB
Algorithms
Keep for microRNA prediction
17
miRDeep2
18
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ?? ????? ?? ?????? ? Bowtie ? Vienna RNA package
• ?????? ????? ?? ????? ??? ???? ?? ? Reads
  ????????, ?? ???? ???? ????, ??????? ?????? ????
  , ??? ?read , ????? ?? ?read ????? ????.
• miRDeep2 package ???? 3 ???????
• 1.miRDeep2 module ????? miRNAs ?????? ??????,
  ????? ????.
• 2.Mapper module ????? ?? ? reads ?????.
• 3.Quantifier module ????? ?? ? reads ?? ? miRNAs
  ???????.

19
?????? ??? ???????
20
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
miRanalyzer mirDeep2
??? Java Perl
Open source ?? ??
????? ?????? ???????? ?? ???? ????
???? ??????? ?? ????
??? ???? ??? ?????
????? ??? txt html, pdf,
????? ????? ????? ?????? ????? ???????
21
???? ??????
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ???? ?? ?? 66 ?????? ???? ???????
• ????? ??????? ?????????? ??? ???
• ????? ????? ??? ???? ??????? ????? ???? ??????
  ????? ???? ??????? ??? TRUE POSITIVE
• ???? ?????? ?? ??????- ????? ?????? ???????
• ????? ???? ??? ???? ????? ?? ???????? ????????
  ???? ??? ???????

22
?????? ??? ???????
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
?????? ??? ?predicted novel miRNAs ????????
??????? miRDeep ?-miRanalyzer.
23
ER ????? ER-
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing
???? ???? ?? ??????? , ?????? ?? ??? ??? ???
??????? ????????. ???? ???? ?? ???? ???? ?????
??????? ???????(ER), ?????? ?????? ?????? ??
??????? ??????? ????? ?????? ??????? ?????
??????? ?? ???? ????? ???.
24
PR ????? PR-
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ???? ???? ?? ??????? ?????? ??????? ??????????,
  ???? ??? ?????? ?????????? ??? ??????, ???????
  ???? ?? ?????? ??????? ??????? ?????????(PR) ,
  ????? ???????? ??????? ?? ?? ???? ?? ????????
  ??????? PR-. ????????? ??? ?????? ??? ??????
  ????? ?? ?????? ????? ?????? ???? ????.
  ?????????? ????? ????? ???? ???? ?????? ?????
  ?????, ?? ????? ????? ?????? , ??????? ???????.

25
HER-2
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• Human Epidermal Growth Factor Receptor ??? ?????
  ???? ????? ( ?????) ????? ????? ???? ????? ??????
  ?? ??? ???? ?? ???? ????? ???. ? HER2 ????? ????
  ????? ?? ?????? ??????? ?? ???? ??????. ??????
  ??? ??? ????? ?????? ?????? ?? ????, ???? ?? ????
  20 ?????? ??????? ??? ????????? ?????? ????? ??
  ????? ????? ??????. ????? ?? ???? ?? ????? ??????
  ????????? ?? ??? ?????? ?????? ???. ???????
  ??????? ???? ????? ?????? ?? ?????? ??????
  HER2-positive. ???? ????? ???? ?? ?????????
  ??????? ????? ?????? ?? HER2 ?????? ????? ?????
  ?????? ?? ???? ??.

26
?????
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ?????, ????? ????? ?? ? miRNAs ???????? ??? ??
  ???????.
• ????? ??? ? miRNAs ???? ?? ???? ??? ?? ?? ??? ??
  ???????, ??"? ????? ??????? ??? ?? ?????????
  ????? miRNAs ???????? ?? ?? ??? ?????.
• ???? ??????? ??????? ????? ???? R, ??????? VENN
  DIAGRAM

27
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• Predicted novel miRNAs ER and ER-

11 predicted novel miRNAs ???????? ?? ???????
ER- ????? ?"? ??? ???????, ??? ???? 46 predicted
novel miRNAs ???????? ?? ??????? ER
28
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• Predicted novel miRNAs PR and PR-

11 predicted novel miRNAs ???????? ?? ???????
PR- ????? ?"? ??? ???????, ??? ???? 18 predicted
novel miRNAs ???????? ?? ??????? PR
29
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• Predicted novel miRNAs HER2 and HER2-

8 predicted novel miRNAs ???????? ?? ???????
HER2- ????? ?"? ??? ???????, ??? ???? 21
predicted novel miRNAs ???????? ?? ??????? HER2
30
?????? ?????....
????? miRNAs ?????? ???? ??? ??????? small RNA
sequencing

• ??????? ?? ????? miRNAs ?????? ??????? ?????
  ?????? ???? ????? ????????, ???????? ??? ??????
  ????? ??????? ????? ???? ??????? ???????? ???????
  ???????. ??????? ??????? ???? ??? miRNAs ???????
  ??? ?? ??? ?? ????? ????? ???, ?miRNAs ?????? ???
  ?? ?????? ??????? ??? ???? ????? , ?????? ???
  ?????? ????? ???? ????? ????? ??????? ?"? ?????
  ??? ? miRNAs ?????? ?????? ?????? ?? ??? ?????.
  ????? ????? ??, ???? ???? ????? ?? ????? ? novel
  miRNAs ??????? ?????? ?? ???? ????? ???? ??? ????
  ???? ???? ?????. miRNAs ??? ?????? ???? ??????? ,
  ?? ????? ???????? ?? ??? ?????, ??? ?????? ??????
  ????? ????? ??????

31

• ???? ?? ??????
• ???? ?
• ?"? ??? ??????
• ????? ???