National Institute of Agricultural Biotechnology (NIAB)

Analysis of the rice gene expression profile by 60K microarrayKim Yong-Hwan1), Lee Jung-Sook,1) Yoon Ung-Han1), Lee Kang-Seob1), Nahm Back-Hie 2), Song Sang-Ik 2),

Choi Sang-Bong 2), Ahn Joong-Hoon 3)

1) National Institute of Agricultural Biotechnology, RDA, Suwon, 441-707, South Korea2) Myoung Ji University, Yong In , 449-728, South Korea3) KonKuk University, Seoul,143-701, South Korea

Rice variety : Dong-Jin Byeo

Rice 60K oligomer chip

- Rice Whole genome Oligomer Chip

- 64,000 spots in two slides

- - Design of 70mers with Gene Prediction

Experimental procedure

- Dye Labeling

- Hybridization

- Two dye analysis ; Cy3 (control) and Cy5 (Treatment)

Microarray analysis:

- Data capturing with GenePix Pro 5.0

- Data processing with Acuity v.3.0

- Statistical Analysis

. Expressed gene profile of rice

- Tissue specific expression (9,768) ; Regeneration tissue ; 790 Flower ; 4,039 Root ; 2,173 Leaf ; 2,766 Transcription factor ; 431

- Two fold change by Hormone treatment (3,287)

Transcription factor ; 249

Future plan To construct the expression gene network in rice, we will used Ac/Ds mutant lines and transgenic rice as many as possible and the obtained DNA chip data will be opened at NIAB home page (NABIC)

ResultsRice 60K Oligomer Microarray

Tissue and stage specific transcriptome profiling with Rice 60K Microarrary

Hierarchical clustering of tissue specific Expression Profiling

Hierarchical clustering of transcription factors Treated with five Plant hormones :

Hierarchical clustering of Expression ProfilingTreated with five Plant hormones

Abstract

Materials and Methods

ConclusionManagement system of gene expression profile of rice at NABIC

Hierarchical clustering of transcription factors at rice tissuesPutative transcription factors in Rice 60K Microarray: 2,528Number of TFs significantly changed between tissue comparison

Hierarchical clustering of significant spots (12,289) were performed.

In order to screen of tissue and plant hormone specific genes from rice, 60K oligomer chips were used. 790 regeneration tissue specific, 4,039 flower specific, 2,173 root specific and 2,766 leaf specific genes were isolated. Among the tissue specific expressed genes, 431 genes were identified as transcription factors. High expressed (2 fold up) 3,287 genes treated by Plant hormones (Jasmonic acid, Salicylic acid, Abscisci acid and Methyl jasmonate) were profiled. 249 out of 3,287 genes were identified as transcription factor. All information of expressed gene of rice DB will be opened at NABIC (National Agricultural Biotechnology Information Center) site (http://www.niab.go.kr )

• 64,904 spots / two slides,

•48 blocks of 12 rows and 4 columns.

•676 (26x26) spots

• 60,100 oligomers

- Known and predicted genes: 57,734

- Antisense : 2,310

- Randomized control DNA : 66

- Blank spots: 4,190

•Average diameter of 120 um.

•70mers with average Tm of 68+/-5° C

•Effective Cost : 10,000 won/gene X 57,000 genes

Reg. callus

Leaf

EtiolatedLeaf

root

FlowerAfter Pollination

callus

Germinating seed

FlowerBeforePollination

Heading

Leaf Heading

Root

Reg.callus

Callus Hd Lf Rt Reg

callus

Hierachial Clustering

Flower stage specific (4,039)

Leaf specific (2,766)Root specific a(2,173)

Root specific b(*)

Reg. Callus specific (790)

Flower specific group (130)

Leaf specific group (110)

Root specific group (93)

Regeneration group (9)

ET JA SA ABAmJAAfter 1 hr

Whole Genes (8000)

ET : 1mM ethephon  (Ethylene) JA : 100mM jasmonic acidSA : 100mM salicylic acidABA : 100mM abscisic acid mJA : 100mM methyl jasmonate

ET JA SA ABAmJA

Transcription Factors (600)After 1 hr

Popup window shows detailed microarray data

Sample, Labeling, Hybridization, Scan information

Gene expression level check

Group 2X Up 2X Down Total

ABA 790 119 909ET 243 282 525JA 1,501 601 2,102mJA 49 46 95 SA 84 258

342

Total 2,284 1,003 3,287

Group 1.6X Up 1.6X Down Total

ERF/AP2 7 10 17BHLH 6 16 22bZIP 3 6 9C2H2 7 9 16C3H 19 17 36Myb 4 8 12Myb-Related 3 4 7NAC 5 8 13WRKY 5 11 16Subtotal 61 92 153Etc 96

Total 249

1.6 fold up regulated transcription factors

Hormone ABA ET JA mJA SA Toatal

TFs

WRKY 3 2 5

ERF 1 5 1 7

NAC 2 3 5

bHLH 1 2 5 2 1 11

Total 5 4 13 5 1 28