1DNARNA

2 Southern blotNorthern blot

3 PCR

PCRPolymerase chain reactionDNARNAcDNA106ugDNADNA

DNADNA3'DNADNADNA4dNTPDNA30-40DNA1pg1ug

PCR

1PCRPCRTaqMg++dNTPC/G

2PCR

3PCRTaqMg++

4

1RACE PCR

2CGAT1:1

3RTPCR

5PCRPCR

6TmPCRTouch Down PCRRACE PCR

7CGDNADMSODIMETHYL SULFOXIDEPCR1/104ulDMSO36ulPCR40ul

8MgCl2PCR bufferMgCl2MgCl2

9PCRTaq10PCR BufferdNTP

1029

11PCRPCRPCRPCRPCR0.5ml

12PCR

PCR

4 PCR

PCRPCR

PCR

PCRPCRTaqPCRSTRATAGENEQuikChange Site-Directed Mutagenesis KitPCR

DNAPfuTurbo DNA polymerasebpDNAPCRPCRDpn IDNADpn IDNAPCRPCR

5 DNA

1DNADDPCR

2

3FootprintDNA

23ddNTP3DNA5DNA3dNTPDNADNA4dNTPddNTPDNADNAddNTPDNA4ddNTPdNTP4ACGTDNA

1DNA

1 DNA 12 ul3-5ug

6 ul

2M NaOH/1 mM EDTA 2 ul

37 10 min

2 7 ul

3M NaAC 6 ul

100 75 ul

5 min

310 min470110ul10 min45 min

4 H2O 6 ul

2 ul

Sequence Buffer 2 ul

65 2 min10-30 min

54 G A T C

Dideoxy Termination Mix ddG ddA ddT ddC

2.5 2.5 2.5 2.5 ul

60.1M DTT 1 ul

Labeling Mix1:4 H2O 2 ul

[35S]dATP 1 ul

Sequenase 2 ul

5 min

7 G A T C

3.5 3.5 3.5 3.5 ul

3710 min

Stop Solution 4 4 4 4 ul

75-803 min

4

2

1

1005ml Repel-silane1000.4mmSpacer

26

5TBE 12 ml

28.8 g

48 ml

40 9 ml

10 0.42 ml

60 ml

TEMED 18 ul

3

DNA

3

1

1TBE1000ml551500V

2

GATCGATC2.5ul2000V

4

1

8080-100

2

XX3

1

2

3

44022

5GATCGATC4GCGC

mRNA

6 RNA

7 mRNA

8 Northern blot

9 RTPCR

10

mRNA

11 cDNA Array Micro Array

cDNA Array cDNA96cDNA

MicroArraycDNA Array

MicroArrayRNAcDNARNA

GeneChip139000216-20mRNA2516-20discoveryARRAYRFDD-PCRPCRMicroArraymRNAPCRmRNAArray

1

2

3RNA

4

5

1

2

3

4

5

6

cDNA ArrayCLONTECH LaboratoriesAtlas cDNA Expression ArrayscDNA ArraycDNA96RNAmRNAmRNAcDNAcDNAArray123

1RNA2Array34Array1200mRNA

ArrayArrayMicroArrayArrayArrayArray

cDNA Array9616000Array

cDNA ArraySouhtern blotNorthern blotcDNA ArrayCLONTECHAtlas cDNA Expression Arrays

CLONTECHDNADNAcDNAmRNAGenBank

1ugmRNAcDNAcDNA ArraycDNA

X

1cDNA ArraysmRNAmRNAmRNA

2RNAmRNAmRNAmRNAmRNA

3[-32P]dATP[-33P]dATP[-32P]dATP[-33P]dATP

4DNAExpressHyb

5

mRNA

12 DDPCR

DDPCR

RNAH-T11-CH-T11-GH-T11-A11TmRNAAmRNAcDNAcDNAAAGCTTGATTGCCPCRcDNA[33P]dATPPCRPCRmRNA

1

Hpa IIpBR 3225GC

pBR3221ul

5RT Buffer4ul

0.1 M DTT0.5ul

[32P]dCTP5ul

2mM dA/T/GTP0.5ul

AMV RT1ul

4230min.

2mM dNTP1ul

4210min.

0.5M EDTA1ul

TE108ul

6510min. 45min.

Sephedex G502000rpm3

100ulpBR 322

cpm1ul3-4105

2DDPCR

1RNAcDNA

0.5mlul

1Z1 2Z2

1 2 3 4 5 6

9.4 9.4 9.4 9.4 9.4 9.4

5RT Buffer 4 4 4 4 4 4

dNTP250uM 1.6 1.6 1.6 1.6 1.6 1.6

RNA0.1ug/ul 22 2 2 2 2

H-T11-M2ul A C G A C G

655min3710min

MMLV 1 1 1 1 1 1

3750min755min

4DDPCR

2DDPCRPCR

1PCR

60

10PCR Buffer 12

dNTP25uM 9.6

-[33P]dATP 1.5

Taq 1.2

20.5ml

1H-AP-1

1-1 1-2 1-3 1-4 1-5 1-6

H-AP-1 2 2 2 2 2 2

H-T11-M2ul A A C C GG

2ulZ1Z2Z1Z2Z1Z2

PCR 14 14 14 14 14 14

18-32

20 20 20 20 20 20

PCR

94 30 s.

40 2 min.

72 30 s.

40

72 5 min.

3

3.5ul2ul802min.X

4DNA

XXDNA300-500bp3MMX

1.5ml100ul302022-5ulPCRPCR40ulDDPCRPCR200ng-1000ngPCR1Northern blot2PCR

GenBankcDNAcDNA

1RNA

2DNARNAGHMessage-Clean kitRNADNAmRNAmRNA

3DDPCR100bp1-2

4DDPCR

5DDPCRNorthern blot

13 cDNA

14 cDNA

15 RACE PCR

RACE PCRRACERapid Amplification of cDNA EndscDNAPCRRACE PCRPCRcDNA

cDNA53cDNAPCRPCRPCRcDNAcDNANorthern blotcDNARACE PCR

1PCR

2123

1PCR23PCRPCRPCRcDNAcDNA

1PCRRACEcDNAcDNAPCRcDNARACE PCR

2Touchdown PCRTouchdown PCRPCRPCR5~10PCR/3~108078767472

3PCRPCR

4Tm7022mers25~30mersC/G45~60%Tm

16 Western blot

17 ELISA

18

19

20

21

GnRH

GnRH

GnRH GnRH

Chart1

20

130

10

240

30

Sheet1

201301024030

Sheet1

Sheet2

Sheet3

22

23 Footprint

24

25

26

27 DNA

28 DNA

29

DNA

30DNA