附錄

附錄一、縮寫對照表

縮寫 全名

1-MCP 1-methylcyclopropene

2,4-D 2,4-dich

aa Amino

ABA Abscisi

ABR ABA-re

ACC 1-amino

AGP Arabino

AmS Ammon

AP Alkalin

APS Ammon

AGP Arabin

AVG Aminoe

BAC Bacteri

BCIP 5-brom

BLA Basic lo

B. or Bipolar

bp Base pair

BPB Bromop

BSA Bovine

C Cystein

CaM Cauliflo

CBB Cooma

CBF C-repea

cDNA Comple

CDS Coding

CHX Cycloheximide

D Asparti

dAT 2’-deoxdCT 2’-deox

- 127 -

lorophenoxy acetic acid

acids

c acid

E sponse element

cyclopropane-1-carboxylic acid

s galgctan proteins

ium sulfate

e phosphatase

ium persulfate

s ogalactan proteins

thoxyvinyl glycine

al artificial chromosome

o-4-chloro-3-indoyl phosphate

ST cal alignment search tool

yzae is oryzae

henol blue

serum albumin

e (Cys)

V35S wer mosaic virus 35S

ssie brilliant blue

t (CRT) binding factors

mentary DNA

sequence

c acid (Asp)

P yadenosine 5’-triphosphate P ycytidine 5’-triphosphate

- 128 -

DEP DiethylC pyrocarbonate

dGT 2’-deox

DMF Dimeth

DMSO Dimeth

DNA Deoxyr

dNT Deoxyn

DRE Dehydr

DTT Dithioth

dTT 2’-deox

E Glutam

EDT Ethylen

EIRE Elicitor

E. co Escheri

Eph Ethephon

ERE Ethylen

ERF Ethylen

EST Express

EtBr Ethidiu

EtOH Ethanol

F Phenyla

FPL Fast pro

G Glycine

GAR Goat-an

GBC Gladiol

GBF G-box b

GCG Genetic

GFP Green f

G. fu ibbere

GlcN β-1,4-N

GRA Reso

HR Hypers

IEF Isoelect

IgG Immunoglobulin

P yguanosine 5’-triphosphate

ylformamide

yl sulfoxide

ibonucleic acid

P ucleoside triphosphate

/CRT ation-responsive element/C-repeat

reitol

P ythymidine 5’-triphosphate

ic acid (Glu)

A ediamine tetra-acetic acid

responsive element

li chia coli

e responsive element

e-responsive transcription factor

ed sequence tag

m bromide

lanine (Phe)

C tein liquid chromatography

(Gly)

ti-rabbit IgG

us bulb class IIIb chitinase

inding factor

s computer group

luorescent protein

jikuroi G lla fujikuroi

Ac -acetylglucosamine

MENE A urce for Comparative Grass Genomics

ensitive response

ric focusing

G

- 129 -

IPTG Isoprop yl-β-D-thiogalactoside

JA Jasmon

K Lysine

kb Kilobase

kDa Kilo Da

LB Luria-bertani

Loc_ Locatio

LTP Lipid-tr

MeJ Methyl

mRN Messen

MS Murash

M. W Molecu

MW Molecu

NBT Nitro bl

NIR Non-im

O. sa Oryza s

Osm Oryza s

Osm Oryza s

PBS Phosph

PCR Polyme

pI Isoelect

PR Pathogenesis-related

PVD Polyvin

Q Glutam

R Arginine

RAG Rabbit-

RA- abbit

RA- abbit

RE Restrict

RIPs Ribosom

RNA Ribonu

rpm Revolu

rRN Ribosom

ic acid

(Lys)

lton

n on chromosomes

s ansfer proteins

A -jasmonic acid

A ger RNA

ige and Skoog

. lar weight

CO lar weight cut off

ue tetrazolium

mune rabbit serum

tiva ativa

30 ativa medium 30-kDa protein

34 ativa medium 34-kDa protein

ate-buffered saline

rase chain reaction

ric point

F ylidene difluoride

ine (gln)

(arg)

-AP anti-goat IgG alkaline phosphatase conjugate

Osm30 R anti-Osm30 antibodies

Osm34 R anti-Osm34 antibodies

ion enzyme

e-inactivating proteins

cleic acid

tions per minute

A al RNA

- 130 -

R. so Rhizoctlani onia solani

RT Room t

SA Salicyli

SAR System

SDS Sodium

SDS odium

SE Standar

SEB Silencin ding factor

S. or Sarocla

SSC Saline-s

TAIR The Ara e

TBC Tulip b

TE Tris-EDTA buffer (10 mM Tris-Cl, pH 8.0, 1 mM EDTA)

TEM N,N,N,

TLPs Thauma teins

TMV Tobacc

Tris Tris (hy ane

UV Ultravio

W Tryptop

W-b WRKY binding site

WRK Plant tr contain WRKY domains

x-gal 5-brom -D-galactopyranoside

Y Tyrosin

emperature

c acid

ic acquired resistance

dodecyl sulphate

-PAGE S dodecyl sulphate -polyacrylamide gel electrophoresis

d err

F g element bin

yzae dium oryzae

odium citrate buffer

bidopsis information resourc

ulb chitinase

ED ’N’-tetramethyl ethylene-diamine

tin-like pro

o mosaic virus

droxymethyl) aminometh

let

han (Trp)

ox

Y anscription factors

o-4-chloro-3-indolyl-β

e (Tyr)

- 131 -

英文 中文

附錄二、中英對照表

Absc 離層isic acid 酸

Alka 鹼性

Apo 胞外

1-am

Chit 幾丁質酶

Cis-acting element 轉錄

Cycl 亞胺

Cyto 細胞

Dele 基因

Elec 電析

Elici 誘引

Embryogenesis 胚胎

Exoc 幾丁

Gluc 醣苷

Glyc 醣脂

Glyc 醣蛋

Glycosyl hydrolase 醣苷

Gree 水母綠螢光蛋白質

Hyp 高張

Hypersensitive response 過敏

Iden 一致

Inclu 包涵

Jasm 茉莉

Lyso 溶菌

Meth 甲基

Microsequencing N端胺基酸微定序

Mor 形態

line phosphatase 磷酸酶

plastic

inocyclopropane-1-carboxylic acid synthase ACC

inase

合成酶

調控位

oheximide 環己酮

kinin 分裂素

tion 缺失

troelution 法

tor 劑

發生

hitinase 質外切酶

anase 酶

olipid 質

oprotein 白

水解酶

n Fluorescent protein

er-osmotic shock 衝擊

反應

tity 度

sion body 體

onic acid 酸

zyme 酶

yl-jasmonate 茉莉酸

phogenesis 發生

- 132 -

Mycorrhizal fungi 菌根菌

N-acetylglucosamine 乙醯葡萄胺

Nitro 硝化

N-gl N-醣

Nylo 尼龍

Osm 滲透

Osm 滲透

Para 寄生

Path 病原

Pathogenesis-related protein 病原

Pept 胜肽

Phyt 植物

Polymerase chain reaction 聚合

Salicylic acid 水楊

Shoo 枝系

Secretory signal peptide 分泌

Simi 相似

Stan 標準

Stress-inducible 逆境

Subc 亞型

Sub- 次亞

Subc 繼代

Syst stance 系統性抗病反應

TMV 菸草鑲嵌病毒

Tran 轉錄

Tran ressor 轉錄

Vector 載體

Zymogram 膠體活性染色

cellulose membrane 纖維膜

ycosylation site 化位

n membrane 膜

otic shock 壓衝擊

otic stress 勢逆境

sitism

ogen 菌

相關蛋白質

idoglycan 聚醣

oalexin 殺菌素

酶鏈反應

酸

ts

訊息序列

larity 度

dard err 差

誘導

lass

subclass 型

ulture 培養

emic acquired resi

s-acting factor 因子

scriptional rep 抑制因子

- 133 -

附錄三、微生物培養基配方

◎ LB medium for bacteria

1 L:

10 g

5 g Yeast extract

5 g NaCl

15 g Agar

- Adjust pH to 7.0 with 3 N NaOH.

- Autoclaved at 121℃ for 15 min.

◎ LB/MgSO4

LB broth (LB medium without agar)

10 mM MgSO4

◎ PDA medium for fungi

1 L:

200 g Potato

- 以電鍋熬煮一小時，隨後以四層紗布過濾，取濾液。

20 g Glucose (dextrose)

20 g Agar

- Autoclaved at 121℃ for 20 min.

Tryptone

- 134 -

養液配方

oncentration in

medium

Stock solution (fold) Stock added

to per liter

附錄四、Murashige-Skoog 培

Stock # Chemicals C

1 2.06 × 10-2 M

1.88 × 10-2 M

82.5 g/L (50 X)

95 g/L (50 X)

20 ml NH4NO3KNO3

2 2O 3.00 × 10-3 M 22.05 g/L (50 X) 20 ml

3 7H2O 1.50 × 10-3 M

18.5 g/L (50 X)

0.754 g/L (50 X)

0.43 g/L (50 X)

20 ml

4 KH2PO4

3

2O

1.25 × 10-3 M

1.00 × 10-4 M

1.00 × 10-6 M

8.5 g/L (50 X)

0.31 g/L (50 X)

41.5 mg/L (50 X)

12.5 mg/L (50 X)

20 ml

5 Na2EDTA⋅2H2O

2 4 2

1.00 × 10-4 M

1.00 × 10-4 M

1.861 g/L (50 X)

1.39 g/L (50 X)

20 ml

6 CuSO4⋅5H2O

2 2

1.00 × 10-7 M

1.00 × 10-7 M

25 mg/L (1000 X)

25 mg/L (1000 X)

1 ml

7 Myo-inositol

e-HCl

mg/L

1 mg/L

10 g/L (1000 X)

1 g/L (1000 X)

1 g/L (1000 X)

1 ml

8 2,4-D 1.00 × 10-5 M 2.21 g/L (1000 X) 1 ml

9 Sucrose 3% (w/v) 30 g

CaCl2⋅2H

MgSO4⋅

MnSO4 9.99 × 10 M

ZnSO4⋅7H2O 2.99 × 10

-5

-5 M

H3BO

KI 5.00 × 10Na2MoO4⋅2H

-6 M

Fe SO ⋅7H O

CoCl ⋅6H O

Nicotinic acid 10

Pyridoxin

Thiamine-HCl 1 mg/L

100 mg/L 100 g/L (1000 X)

- Adjust pH to 5.8 with 3 N NaOH.

- Autoclaved at 121℃ for 15 min.

- Stocks 1-6 are stored at 4℃ and stocks 7-8 at -20℃. Sucrose is added freshly.

- This recipe is originated from Dr. L.-F. Liu, Dep. of Agronomy, NTU.

- 135 -

s r

Final concentratio Stocks V

附錄五、RNA 電泳所使用的藥品配方

◎ RNA ample buffe

n olume (μl)

50% (v/v) formamid 10 1e 0% 25

10% (v/v) formaldeh 37 67.5

4, pH 6 1 1

0. 5

l EtBr 10 5

A 37.5

ume (μl) 250

yde %

40 mM NaPO .8 M, pH 6.0 0

10 mM EDTA 5 M

0.2 mg/m mg/ml

DEPC-H2O utoclaved

Total vol

◎ 5X RNA loading buffer

n S Volume (μl)Final concentratio tocks

50% (v/v) glycerol 80 3% 12.5

5 mM NaPO4, pH 6. 1M 2.5

moph 1% 5

lene c 1% 50

DEPC-H 85

l) 500

8 , pH 6.0

0.1% (w/v) bro enol blue 0

0.1% (w/v) xy yanol

2O Autoclaved

Total volume (μ

◎ RNA g er

ncentration S V (ml)

el running buff

Final co tocks olume

10 mM NaPO4, pH 6 1M, pH 6.0 1.8 0

8% formaldehyde 37% 216

Autoclaved 774

1000

DEPC-H2O

Total volume (μl)

- 136 -

irection* RE Sequence

附錄六、PCR primers

Primers D

Osm

(Osm

Bam

HI GGA TCC30-1 F

30-F) 5’-CTC ATG GCT GCT AAT

AAG CTC-3’

Osm Bam

HI GGA TCC30-2 F 5’-TAT GGC GAC ATC GCC

Osm

(Osm

R Hind

III AAG CTT

GTG TAC-3’

30-3

30-R) 5’-CTC GAC GCT GCT CTT

-3’

Osm

(P1)

F Bam

HI TCC

CAC CTG

34-1 5’-TTA GGA ATG GGC TCC GCC

AAG-3’

Osm34-3 F Bam

HI 5’-CTC GGA TCC

(P2) GTA AAC TCC AAC

CTC-3’

Osm Hind

III AAG CTT34-4 R 5’-CTC GTA A

-3’

Poly Xba

I TCTAGA

CT GAT GGT

GGC

T (P3) R 5’-GTCGAC

TTTTTTTTTTTTT-3

GFP-1 F Bam

HI C AGG GAT CC

TT ’

5’- G AGT AAA GGA

GFP-2 R Hind

III 5’-CTC AAG CTT

GAA GA-3’

GTA TAG TTC ATC

CAT G-3’’

OsmChi1-5 F Eco

RI 5’-TGA GAATTC GAG CAG TGC GGC

AG-3’

Osm Bam

HI GGATCCChi1-7 F 5’-AAT GAG C

Osm Hind

III TC AAG CTT

AG TGC GGC

AG-3’

Chi1-6 R 5’-C CCT C T GTA TG GT

GCA-3’

註：「F」為於目標基因 5’端的正向 (forward) 引子，即為 sense primer；「R」

為於目標基因 3’端的反向 (reverse) 引子，即為 antisense primer。

*

- 137 -

第三型幾丁質酶 Osm34 的序列資料

LOCUS 0230 941 mRNA linear N-2003

DEFINITION Oryza sativa chitinase mRNA, partial cds.

ACCESSION AF330230

VERSI 0.1 GI:27462187

KEYWO

SOURCE a sativa (ja ica cultivar oup)

ORGANISM Oryza sativa (japonica cultiv roup

aryota; Viri lantae; St hyta

ophyta; Magnoliophyta; iops Poaceae; BEP

de; Ehrharto ae; Oryzea za.

REFEREN bases 1 to 9

AUT Li,C.-W. and Wang,Y.-C.

TIT tial cDNA se nce encodi hiti

suspension-cultured rice cell

JOU ublished

REFEREN bases 1 to 9

AUTHORS Li,C.-W. and Wang,Y.-C.

TIT t Submissi

JOURNAL Submitted (19-DEC-2000) Depart nt of n

ormal Universi 88 Ting-C ., Sec. 4, Taipei, Taiwan 11718,

epublic of Chin

FEATURES Location/Qualifiers

1..941

/organism="Oryza sat (jap ar-group)"

/mol_t ="mRNA"

/strain N5"

/db_xref="taxon:3994

- 138 -

caacatctt ctggcagaac

ggc cgtcgcgtcg atcaagcaga gcaaccccaa cgtgagggtg

tgaac gaccgcccgg tcttcttcaa catcacctcc

ctcgt gggtaaacaa cgctgtcgag tccctcaccg gcatcatcca ggacaacaac

g acccggacac cttcaccgag

g tgatcaagtt cgcctccatc

cc ccggagggaa catccttgcg

acgacctcg gtgcccgtcg agacggcgct gagcgcttgc

tgtac ggcatcttca tctgggctgc tgatcattcc

ccatc

attattg tttcacattt ttaaacaaag

actatg aaaataagca aaaaaaaaaa a

ORIGIN

1 gtaaactcca acctcttccg ggactacatc ggcgccatct tcaacggcgt gaagttcacc

61 gacgtgccaa tcaatccgaa agtccggttc gacttcatcc tcgccttcat catcgactac

121 accaccgaga cgaacccgcc gaccccgacc aacggcaagt t

181 acggtgctca ccccgtc

241 gccgtcagca tgggcggcgc caccg

301 gtcga

361 ctggacggca tcgacatcga ctacgagcag ttccaggtc

421 tgcgtcggcc gcctcatcac ggtgctcaag gccaagggg

481 gcgcccttcg gcaacgccga ggtccagcgc cattacatgg cgctgtgggc caagtacggc

541 gccgtgatcg actacatcaa cttccagttc tacgcctacg gcgccagcac gacggaggcg

601 cagtacgtcg acttcttcaa ccagcagatc gtcaacta

661 agcttcacga cggcggcgac g

721 cggactctgc agaaggaggg gaagc

781 aggagccaag ggttcaagta tgagacagaa tcgcaagcac ttctggctaa tgcca

841 agttactagc tagtagctag ttttgtttgc tgc

901 atcacaagag ctgt

//

- 139 -

OsmChiI-34

URNAL Unpublished

EFERENCE 2 (bases 1 to 894)

AUTHORS Li,C.-W., Li,S.-H. and Wang,Y.-C.

TITLE Direct Submission

JOURNAL Submitted (02-SEP-2003) Department of Life Science, National Taiwan

Normal University, 88 Ting-Chou Rd., Sec. 4, Taipei, Taiwan 116,

Republic of China

FEATURES Location/Qualifiers

source 1..894

/organism="Oryza sativa (japonica cultivar-group)"

/mol_type="mRNA"

/cultivar="Tainan 5"

/db_xref="taxon:39947"

CDS 894

/EC_number="3.2.1.14"

/note="class I chitinase; secreted by suspension-cultured

rice cells"

/codon_start=1

/product="OsmChiI-34"

/protein_id="AAQ84333.1"

/db_xref="GI:35187685"

附錄八、水稻細胞分泌之第一型幾丁質酶 的序列資料

LOCUS AY378172 894 bp mRNA linear PLN 29-SEP-2003

DEFINITION Oryza sativa (japonica cultivar-group) OsmChiI-34 mRNA, partial

cds.

ACCESSION AY378172

VERSION AY378172.1 GI:35187684

KEYWORDS .

SOURCE Oryza sativa (japonica cultivar-group)

ORGANISM Oryza sativa (japonica cultivar-group)

Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta;

Spermatophyta; Magnoliophyta; Liliopsida; Poales; Poaceae; BEP

clade; Ehrhartoideae; Oryzeae; Oryza.

REFERENCE 1 (bases 1 to 894)

AUTHORS Li,C.-W., Li,S.-H. and Wang,Y.-C.

TITLE Partial cDNA sequence encoding a class I chitinase secreted by

suspension-cultured rice cells

JO

R

- 140 -

PQSPKPSCHAVMTGQWTPNGNDQAAGRVPGYGVVTNIINGGVECGHGADSRVADRIGF

LGVSYGANLDCYNQR"

gtgcg gcagccaggc cggcggcgcg ctctgcccca actgcctctg ctgcagccag

g gctgccagag ccagtgctcc

g gtggcggcag cggcgtcgcc

gg tcgccgcgtt cctggcgcag

ggctggccc acggcgcccg acggccccta ctcctggggc

acat actgcgagcc caagccggag

tccaacc tgctgagcaa cccggacctg

caccgt ctccctcaag acggcgttct ggttctggat gaccccgcag

gcggtgatg acagggcagt ggacgcccaa cggcaacgac

acca acatcatcaa cggcggcgtg

cagcagagtc gccgaccgga ttgggttcta caagcgctac

/translation="EQCGSQAGGALCPNCLCCSQYGWCGSTSDYCGSGCQSQCSGSCG

GGGPTPPSGGGGSGVASIVSRSLFDQMLLHRNDAACPAKNFYTYDAFVAAANAFPSFA

TTGDAATRKREVAAFLAQTSHETTGGWPTAPDGPYSWGYCFKEENNGNAPTYCEPKPE

WPCAAGKKYYGRGPIQITYNYSYGPAGQAIGSNLLSNPDLVASDATVSLKTAFWFWMT

YKRYCDM

ORIGIN

1 gagca

61 tacggctggt gcggctccac ctccgattac tgcggctcc

121 ggcagctgcg gcggcggcgg cccgaccccg ccctccggc

181 tccatcgtgt cgcgctcgct cttcgaccag atgcttctcc accgcaacga cgcggcgtgc

241 ccggccaaga acttctacac ctacgacgcc ttcgtcgccg ccgccaacgc cttcccgagc

301 ttcgccacaa ccggcgacgc cgccacccgc aagcgcga

361 acgtcccacg agaccaccgg c

421 tactgcttca aggaggagaa caacggcaac gccccc

481 tggccgtgcg ccgccggcaa gaagtactac ggccggggac ccatccagat cacctacaac

541 tacagctacg gcccggcggg gcaggccatc ggc

601 gtggcgtcgg acgc

661 tcgcccaagc cgtcgtgcca c

721 caggcggcgg ggcgcgtacc gggctacggc gtggtg

781 gagtgcggcc acggcgcgga

841 tgcgacatgt tgggcgtcag ctacggcgcc aacttggact gctacaacca gagg

//

- 141 -

Osm34/pBS Osm34/pET24a 附錄九、 和 圖譜

附錄十、Osm30/pBS 和 Osm30/pET24a 圖譜

- 142 -

- 143 -

圖譜 附錄十一、GFP 的核苷酸序列與 GFP/pET24b

GFP CDS sequence from pCAMBIA1302

Start GFP-1 primer

TG AGT AAA GGA GAA GAA A CTT TTC ACT GGA GTT GTC CCA ATT CTT

TT

T

AAC

GTT GAA TTA GAT GGT GAT GTT AAT GGG CAC AAA TTT TCT GTC AGT GGA GAG GGT GAA GGT GAT GCA ACA TAC GGA AAA CTT ACC CAAA TTT ATT TGC ACT ACT GGA AAA CTA CCT GTT CCA TGG CCA ACA CTT GTC ACT ACT TTC TCT TAT GGT GTT CAA TGC TTT TCA AGA TAC CCA GAT CAT ATG AAA CAG CAT GAC TTT TTC AAG AGT GCC ATG CCC GAA GGT TAT GTA CAG GAA AGA ACT ATA TTT TTC AAA GAT GAC GGG AAC TAC AAG ACA CGT GCT GAA GTC AAG TTT GAA GGT GAT ACC CTTGTT AAT AGA ATC GAG TTA AAA GGT ATT GAT TTT AAA GAA GAT GGA AAC ATT CTT GGA CAC AAA TTG GAA TAC AAC TAT AAC TCA CAC AATGTA TAC ATC ATG GCA GAC AAA CAA AAG AAT GGA ATC AAA GT

TTC AAA ATT AGA CAC AAC ATT GAA GAT GGA AGC GTT CAA CTA GCA GAC CAT TAT CAA CAA AAT ACT CCA ATT GGC GAT GGC CCT GTCCTT TTA CCA GAC AAC CAT TAC CTG TCC ACA CAA TCT GCC CTT TCG AAA GAT CCC AAC GAA AAG AGA GAC CAC ATG GTC CTT CTT GAG TTT GTA ACA GCT GCT GGG ATT ACA CAT GGC ATG GAT GAA CTA TAC AAA TAA GFP-2 primer Stop

- 144 -

生長曲線

A. E. coli

附錄十二、細菌轉形 Osm30、Osm34 和 GFP 基因後的

XL1-Blue strain

Incubation time (h)

0

20

40

60

80

100

Rel

ativ

e am

ount

(%)

pBSpBS-Osm30 pBS-Osm34pBS- GFP

0 5 10 15 20 25

B. E. coli BL21 (DE3) strain

Incubation time (h)

0 5 10 15 20 25

Rel

ativ

e am

ount

(%)

0

20

40

60

80

100

pET24a-Osm30pET24a-Osm34 (307-25)pET24b-GFP

附錄十三、Osm30 基因啟動子序列

- 145 -

附錄十四、Osm34 基因啟動子序列

- 146 -

附錄附錄一、縮寫對照表 附錄二、中英對照表 附錄三、微生物培養基配方 附錄四、Murashige-Skoog培養液配方 附錄五、RNA電泳所使用的藥品配方 附錄六、PCR primers 附錄七、水稻細胞分泌之第三型幾丁質酶Osm34的序列資料 附錄八、水稻細胞分泌之第一型幾丁質酶OsmChiI-34的序列資料 附錄九、Osm34/pBS 和Osm34/pET24a圖譜 附錄十、Osm30/pBS 和Osm30/pET24a圖譜 附錄十一、GFP的核苷酸序列與GFP/pET24b圖譜 附錄十二、細菌轉形Osm30、Osm34和GFP基因後的生長曲線 附錄十三、Osm30基因啟動子序列 附錄十四、Osm34基因啟動子序列