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
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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
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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
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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
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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)
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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
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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
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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
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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)
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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。
*
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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
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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