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3 DNA A Novel DNA Method to Identify & Quantify Plant Materials Summary PCR We provide the simple PCR based technology to identify and quantify the plant species or varieties used as raw materials in foods or herbal medicine products. Features of technology DNA (Species) PCR PCR DNA Achievements PCR PCR

DNA 2 3 5 6 M 2 3 5 6 PCR : 2 : 3 : : A 5 : B 6 : PCR PCR Species 2 3 5 6 7 8 9 2 3 5 6 7 8 9 2 2 22 23 2 25 26 27 28 29 3 3 32 33 3 35 36 37 38 39 2 3 5 6 7 8 9 5 5 52 53 5 55 56 57 58 59 6 6 62 63

4 DNA M PCR : 2 : 3 : : A 5 : B 6 : PCR PCR Species () A B C D E F G H I J K LM N O P Q R S T U V W Recent development of the technology Rtsp-659 Tpv292 PCR PCR PCR GAIIx Rtsp-: MOL GEN & GEN.Vol 272, 6-27, DOI:.7/s Tpv2: PLT MOL BIOL Vol 39, , DOI:.23/A: Prospect of the technology 23 DNA PCR PCR LAMP 65 LAMP (Loop-Mediated Isothermal Amplification): SARS TEL: tahara@cc.okayama-u.ac.jp LAMP DNA DNA DNA

5 ..35 FS-2 Fresh weight(g)/seedling control None FS-2 Fsi AF5553 Penicillium islandicum isola... AY78785 Penicillium rugulosum strain... AY Penicillium variabile strain... DQ Penicillium piceum strain AT... AB3539 Penicillium marneffei rev AF559 Penicillium chrysogenum isol... NW Aspergillus oryzae RIB... OSH A AY Talaromyces wortmannii isola... Length of roots (mm) ppm β-caryophyllene

6 ( n=6; *, P.5) β-caryophyllene (ppm) 8 6 (***, P.) Sowing seeds Exposed to caryophyllene Measured weight Inoculated Observed infection Lesion (days) β-caryophyllene(ppm) TEL: tomoshir@cc.okayama.u.ac.jp

7 Organic Compounds that Specifically Inhibit Chitinolytic Enzymes HOH 2 C O n HO HO OH NHAc N- (GlcNAc) -N- (GlcNAcase) Streptomyces anulatus NBRC3369 TMG-chitotriomycin* Pochonia suchlasporia var. suchlasporia Pochonicine** *Hirokazu Usuki et.al., J. Am. Chem. Soc., 3, 6-52 (28) **Hirokazu Usuki et.al., Bioorg. Med. Chem.,7, (29) GlcNAcase () (2) (3) NagZ TMG-chitotriomycinPochonicine TMG-chitotriomycin () GlcNAcases Plant Mammal Fungus Insect Pochonicine () TMG-chitotriomycin GlcNAcase

Streptomyces anulatus NBRC3369 (TMG-chitotriomycin) TMG-chitotriomycinGlcNAcase TMG-chitotriomycin TMG-chitotriomycin ph 95 C, 5 min 3 7 TMG-chitotriomycin 83, 7~%, 5~7% TMG-chitotriomycin

8 Streptomyces anulatus NBRC3369 (TMG-chitotriomycin) TMG-chitotriomycinGlcNAcase TMG-chitotriomycin TMG-chitotriomycin ph 95 C, 5 min 3 7 TMG-chitotriomycin 83, 7~%, 5~7% TMG-chitotriomycin TMG-chitotriomycin A 2 3 MeOH 3 º C, 6 h chitotriomycin TMG- A tr (min) 3 5 tr (min) 6 MeOH TMG-chitotriomycin!! TMG-chitotriomycin!? Pochonia suchlasporia var. suchlasporia (Pochonicine) Pochonicinepochonicinepochonicine MeOHGC-MS PochonicineA,B R 2 R 3 Pochonicine PochonicineC MeOH 5 R R ~R 3 : OH (2) : H () Pochonia suchlasporia var. suchlasporia () A B C Pochonicinepochonicine tr (min) Pochonicine Pochonicine Pochonicine Pochonicine pochonicinepochonicine : TEL : nitoda@cc.okayama-u.ac.jp

ኚ ຊ䜢䛧䛯ᇵ㣴䠞䜢䛔䛯᪂つ䛺ᢠయぶ ᛶᡂ 䝅䝇䝔䝮 Novel system for affinity maturation of monoclonal antibodies using the

vitroᢠయస〇䝅䝇䝔䝮 ព䛾ᢠయ ኚ㒊䜢DT䛻ᑟධ䛧䚸DTෆ䛷ኚ 䛸㑅ᢥ䛾䜚䛧䛻䜘䜚ぶ ᛶᡂ DT䛾ᐃᖖ㒊䜢䝠䝖IgGᆺ 䛧䚸䝠䝖ᆺᢠయ䛸䛧䛶ᢠయస〇䛜㻰㼀㻠㻜㻙㻿㼃䜢

ᐶᐜ䜢ᅇ㑊䛧䛯ᢠయ䝷䜲䝤䝷䝸䞊䜢 స〇䛷䛝䜛䚹㻌 స〇䛷䛝䜛䚹 ᢠయ䜢䝷䜲䝤䝷䝸䞊䛛䜙䛾ᢠయ ᚓ䛿㻞㻟㐌㛫䛷䛩䜛䚹㻌䛩䜛䚹 ኚ 䛸㑅ᢥ䛾䝃䜲䜽䝹䜢䜚䛧䛶ᢠయ䛾ぶ ᛶ䜢 ᨵⰋ䛷䛝䜛䚹㻌 ᨵⰋ䛷䛝䜛䚹

㻰㼀㻠㻜㻙㻿㼃ෆ䛷ኚ 䛥䛫䚸ᶵ ᨵⰋ䛩䜛䛣䛸䛜䚹㻌㻰㼀㻠㻜㻙㻿㼃ෆ䛷ኚ 䛥䛫䚸ᶵ ᨵⰋ䛩䜛䛣䛸䛜䚹䠄䠏䠅䝠䝖ᆺᢠయ䛾 ᚓ㻌䠄䠏䠅䝠䝖ᆺᢠయ䛾 ᚓ 㻰㼀㻠㻜㻙㻿㼃䜢䝠䝖㻵㼓㻳㻝ᆺᢠయ ᆺ䛻ᨵኚ䛧䛶䛚䜚䚸 ኚ

䚸䝠䝖ᆺᢠయ䠚 ᗘ䛾ኚ 䛸㑅ᢥ䛻䜘䜛ぶ ᛶᡂ 㻌ぶ ᛶᡂ ኚ ኚ ON䛷ᇵ㣴㑅ᢥ ච ᐶᐜ䛾ไ㝈䜢ᅇ㑊䛧䛯ᢠయ 䝷䜲䝤䝷䝸䞊䛾ᙧᡂ㻌㻔㻝㻜䝷䜲䝤䝷䝸䞊䛾ᙧᡂ 㻔㻝㻜㻤䠅䠅㻌 ኚ 䝟䝍䞊䞁䛾ษ

ON -䝠䝗䝻䜻䝅䝍䝰䜻䝅䝣䜵䞁 IgV g loxp 㧗ぶ ᛶᢠయ 㧗ぶ ᛶᢠయ㻌 mutations OFF ON 㻯㼞㼑㻙㼘㼛㼤㻼ࢩࢫ 㻭㻵㻰㑇ఏᏊ䛾㻭㻵㻰㑇ఏᏊ䛾㻌㼄㻾㻯㻯㻟㻌㻗㻛㻙㻌㼄㻾㻯㻯㻟

5 GFP (AID) 2䞄㛫ᇵ㣴ᚋ Ᏻᐃ 㼜㼛㼕㼚㼠㻌㼙㼡㼠㼍㼠㼕㼛㼚㻌㼜㼛㼕㼚㼠㻌㼙㼡㼠㼍㼠㼕㼛㼚㻵㼓㼂㻌㼓㼑㼚㼑㻌㻵㼓㼂㼓㼑㼚㼑 ኚ 䝟䝍䞊䞁䛾ษ䜚䛘䛻䜘䜛㻌 ኚ 䝟䝍䞊䞁䛾ษ䜚䛘䛻䜘䜛 ᢠཎ

6 㼓㼑㼚㼑㻌㼏㼛㼚㼢㼑㼞㼟㼕㼛㼚㻌㻔㼄㻾㻯㻯㻟౫Ꮡ䠅㼓㼑㼚㼑㻌㼏㼛㼚㼢㼑㼞㼟㼕㼛㼚㻌㻔㼄㻾㻯㻯㻟౫Ꮡ䠅㻌 ψ 㼂㻌㼂 IRES ኚ ᶵ 䜢㻻㻲㻲䛻䛩䜛䛣䛸䛻䜘䜛㻌 ኚ ᶵ 䜢㻻㻲㻲䛻䛩䜛䛣䛸䛻䜘䜛 ᢠཎ ᢠయ䛾

9 ኚ ຊ䜢䛧䛯ᇵ㣴䠞䜢䛔䛯᪂つ䛺ᢠయぶ ᛶᡂ 䝅䝇䝔䝮 Novel system for affinity maturation of monoclonal antibodies using the hypermutating B cell line ᒸᒣ Ꮫ Ꮫ㝔 Ꮫ 㻌㔠ᒣ㻌 ᶞ䚸㻌 ṇᶞ䚸㻌㻌㻌 ᇶ ᢏ 䛾ᴫせ ᢠయ㑇ఏᏊ䜈䛾ኚ ຊ䜢䛩䜛䝙䝽䝖䝸B DT䜢䛔䛶᪂つ䛺in vitroᢠయస〇䝅䝇䝔䝮 ព䛾ᢠయ ኚ㒊䜢DT䛻ᑟධ䛧䚸DTෆ䛷ኚ 䛸㑅ᢥ䛾䜚䛧䛻䜘䜚ぶ ᛶᡂ DT䛾ᐃᖖ㒊䜢䝠䝖IgGᆺ 䛧䚸䝠䝖ᆺᢠయ䛸䛧䛶ᢠయస〇䛜㻰㼀㻠㻜㻙㻿㼃䜢䛔䛯㼕㼚㻌㼢㼕㼠㼞㼛ᢠయస〇䝅䝇䝔䝮䛾ཎ 㻌䠘㻰㼀㻠㻜䌦㻿㼃䝅䝇䝔䝮䛾 ᚩ䚸 ᛶ䠚㻌䠘㻰㼀㻠㻜䌦㻿㼃䝅䝇䝔䝮䛾 ᚩ䚸 ᛶ䠚 DT-SW 㻌䠄䠍䠅᪂つᢠయ䛾᥈ 㻌䠄䠍䠅᪂つᢠయ䛾᥈ ᇵ㣴䛰䛡䛷ච ᐶᐜ䜢ᅇ㑊䛧䛯ᢠయ䝷䜲䝤䝷䝸䞊䜢 స〇䛷䛝䜛䚹㻌 స〇䛷䛝䜛䚹 ᢠయ䜢䝷䜲䝤䝷䝸䞊䛛䜙䛾ᢠయ ᚓ䛿㻞㻟㐌㛫䛷䛩䜛䚹㻌䛩䜛䚹 ኚ 䛸㑅ᢥ䛾䝃䜲䜽䝹䜢䜚䛧䛶ᢠయ䛾ぶ ᛶ䜢 ᨵⰋ䛷䛝䜛䚹㻌 ᨵⰋ䛷䛝䜛䚹 ༢㞳䛧䛯ᢠయ䛿䚸ኚ ᶵ 䜢㻻㻲㻲䛻䛧䛶ኚ 䛜ධ䜙䛺䛔䜘䛖䛻Ᏻᐃ 䛷䛝䜛䚹㻌䛺䛔䜘䛖䛻Ᏻᐃ 䛷䛝䜛䚹䠄䠎䠅 ᢠయ䛾ᶵ ᨵኚ㻌䠄䠎䠅 ᢠయ䛾ᶵ ᨵኚ ᢠయ እ㒊䛛䜙䝠䝖䛺䛹䛾ᢠయ ኚ㒊㑇ఏᏊ䜢ᑟධ䛧䚸䛾༢㞳㻰㼀㻠㻜㻙㻿㼃ෆ䛷ኚ 䛥䛫䚸ᶵ ᨵⰋ䛩䜛䛣䛸䛜䚹㻌㻰㼀㻠㻜㻙㻿㼃ෆ䛷ኚ 䛥䛫䚸ᶵ ᨵⰋ䛩䜛䛣䛸䛜䚹䠄䠏䠅䝠䝖ᆺᢠయ䛾 ᚓ㻌䠄䠏䠅䝠䝖ᆺᢠయ䛾 ᚓ 㻰㼀㻠㻜㻙㻿㼃䜢䝠䝖㻵㼓㻳㻝ᆺᢠయ ᆺ䛻ᨵኚ䛧䛶䛚䜚䚸 ኚ 㻻㻲㻲䛻䛧䛶 ᚓᢠయ䛿䝠䝖㻵㼓㻳㻝䛸䛧䛶 άᛶ䜢ホ౯ 䚹㻌 ᚓᢠయ䛿䝠䝖㻵㼓㻳㻝䛸䛧䛶 άᛶ䜢ホ౯ 䚹 Ᏻᐃ Ᏻᐃ 㻌㻌䠘 ᛶ䚸ච ᐶᐜ䛾ᅇ㑊䚸ぶ ᛶᡂ 䚸䝠䝖ᆺᢠయ䠚㻌䠘 ᛶ䚸ච ᐶᐜ䛾ᅇ㑊䚸ぶ ᛶᡂ 䚸䝠䝖ᆺᢠయ䠚 ᗘ䛾ኚ 䛸㑅ᢥ䛻䜘䜛ぶ ᛶᡂ 㻌ぶ ᛶᡂ ኚ ኚ ON䛷ᇵ㣴㑅ᢥ ච ᐶᐜ䛾ไ㝈䜢ᅇ㑊䛧䛯ᢠయ 䝷䜲䝤䝷䝸䞊䛾ᙧᡂ㻌㻔㻝㻜䝷䜲䝤䝷䝸䞊䛾ᙧᡂ 㻔㻝㻜㻤䠅䠅㻌 ኚ 䝟䝍䞊䞁䛾ษ 䛻䜘䜛㧗ぶ ᛶᢠయ䛾 ᚓ㻌䝟䝍䞊䞁䞁䛾ษ 䛻䜘䜛㧗ぶ ᛶᢠయ䛾 ᚓ ኚ ᑟධ䜢㻻㻺㻛㻻㻲㻲䛷䛝䜛䝙䝽䝖䝸㻮㻰㼀㻠㻜㻙㻿㼃㻌 ኚ ᑟධ䜢㻻㻺㻛㻻㻲㻲䛷䛝䜛䝙䝽䝖䝸㻮㻰㼀㻠㻜㻙㻿㻿㼃 ኚ ኚ OFF ኚ ON -䝠䝗䝻䜻䝅䝍䝰䜻䝅䝣䜵䞁 IgV g loxp 㧗ぶ ᛶᢠయ 㧗ぶ ᛶᢠయ㻌 mutations OFF ON 㻯㼞㼑㻙㼘㼛㼤㻼ࢩࢫ 㻭㻵㻰㑇ఏᏊ䛾㻭㻵㻰㑇ఏᏊ䛾㻌㼄㻾㻯㻯㻟㻌㻗㻛㻙㻌㼄㻾㻯㻯㻟㻗㻛㻙 GFP+ Puromycin resistant ෆᅉᛶAID㑇ఏᏊ䜢䛣䛾䝁䞁䝇䝖䝷䜽䝖䛷 AID CAGp loxp GFP P Puror ኚ 㻻㻲㻲㻌 ኚ 㻻㻲㻲 Ᏻᐃ GFP (AID) 2䞄㛫ᇵ㣴ᚋ Ᏻᐃ 㼜㼛㼕㼚㼠㻌㼙㼡㼠㼍㼠㼕㼛㼚㻌㼜㼛㼕㼚㼠㻌㼙㼡㼠㼍㼠㼕㼛㼚㻵㼓㼂㻌㼓㼑㼚㼑㻌㻵㼓㼂㼓㼑㼚㼑 ኚ 䝟䝍䞊䞁䛾ษ䜚䛘䛻䜘䜛㻌 ኚ 䝟䝍䞊䞁䛾ษ䜚䛘䛻䜘䜛 ᢠཎ ᢠయ䛾ぶ ᛶᡂ 䛾㻌 ᢠཎ ᢠయ䛾ぶ ᛶᡂ 䛾䛾ぶ ᛶᡂ 䛾 NP-specific clone XRCC3 (+/+), XRCC3 (+/ ) (+/ / ) (AID-OFF) ኚ 㻻㻺 ኚ 㻻㻺㻌 98.6 㼓㼑㼚㼑㻌㼏㼛㼚㼢㼑㼞㼟㼕㼛㼚㻌㻔㼄㻾㻯㻯㻟౫Ꮡ䠅㼓㼑㼚㼑㻌㼏㼛㼚㼢㼑㼞㼟㼕㼛㼚㻌㻔㼄㻾㻯㻯㻟౫Ꮡ䠅㻌 ψ 㼂㻌㼂 IRES ኚ ᶵ 䜢㻻㻲㻲䛻䛩䜛䛣䛸䛻䜘䜛㻌 ኚ ᶵ 䜢㻻㻲㻲䛻䛩䜛䛣䛸䛻䜘䜛 ᢠཎ ᢠయ䛾 ᛶ䛾ᅛᐃ㻌 ᢠཎ ᢠయ䛾 ᛶ䛾ᅛᐃ NP-BSA binding ᛶ䛜ᑠ䛥䛟ኚ ᛶ䛜ᑠ䛥䛟ኚ 㻌䠄ぶ ᛶᡂ 䠅㻌䠄ぶ ᛶᡂ 䠅 ᢠయ ᢠయ㻌 Mer-Cre-Mer 䛾άᛶ AID P ᛶ䛜䛝䛟ኚ ᛶ䛜䛝䛟ኚ 㻌䠄䝷䜲䝤䝷䝸䞊స〇䠅㻌䠄䝷䜲䝤䝷䝸䞊స〇䠅 Culture for 6 weeks (AID-ON) Single cell e sorting

ኚ ຊ䜢䛧䛯ᇵ㣴䠞䜢䛔䛯᪂つ䛺ᢠయぶ ᛶᡂ 䝅䝇䝔䝮㻰㼀㻠㻜㻙㻿㼃䛻䜘䜛 ᢠయ䛾ᶵ ᨵኚ㻔䝙䝽䝖䝸ᢠయᆺ㻕㻌㼂㻴㑇ఏᏊ㻌㑇ఏᏊ༢㞳㼂㻴㑇ఏᏊ ᛶ䜔ぶ ᛶ䛾 ᨵⰋ䛥䜜䛯mAb 㼂㻸㑇ఏᏊ㻌㼂㻸㑇ఏᏊ ᶵ ᨵኚ䛧䛯䛔 mab(䝠䝖䚸䝬䜴䝇) 䐟ᢠయ㑇ఏᏊᗙ䜈䛾㒊㑇ఏᏊᑟධ ኚ㒊 ᐃᖖ㒊 ᢠయ㑇ఏᏊᗙ እ ኚ㒊 እ᮶ᢠయ ኚ㒊䝍䞊䝀䝔䜱䞁䜾䝧䜽䝍䞊䝬䞊䜹䞊 loxp 䛘 ኚ ON

3 2 2 % 3 2 2 3 <APC-A> 5 NP binding <PE-A> 3 䝬䜴䝇VH 2 FL-H 3 Sorting 䐡DT-SW䛻䜘䜛䝬䜴䝇mAb䛾ぶ ᛶᡂ 䐡 DT-SW DT SW䛻䛻䜘䜛䝬䜴䝇mAb mab䛾䛾ぶ ᛶᡂ ぶ ᛶ䛜 Relative affinity y 㒊㒊㑇ఏᏊᑟධ 5 3 3 2 3 <APC-A> Sorting 2 FL-H 3 2

% 2 2 2 3 2 2 <PE-A> FL2-H FL2-H 䝬䜴䝇Vλ ᐃ ᯝ 䝠䝖ᆺ㻰㼀㻠㻜㻙㻿㼃䝅䝇䝔䝮㻌㻯㻴㻦䝠䝖㻯γ 㻝㻝㻌㻯㻴㻌㻦㻌䝠䝖㻯γ 㻯㻸㻦䝠䝖㻯κ 㻌㻯㻸㻌㻦㻌䝠䝖㻯κ 䜈䛾㻌䜈䛾㻰㼀㻠㻜㻙㻿㼃㻌㻰㼀㻠㻜㻙㻿㼃㻰㼀㻠㻜㻙㻿㼃㻙㼔㼓㻌㻰㼀㻠㻜㻙㻿㼃㻙㼔㼓㼓 ኚ 㻻㻺㻌 ኚ 㻻㻺㼂㻴㻘㼂㻸䛾㻌㻌㻜㻚㻡㻜㻚㻠

䜽䝻䞊䞁䛾䝇䜽䝸䞊䝙䞁䜾㻺㻼㼎㼕㼚㼐㼕㼚㼓㻺㻼㻌㼎㼕㼚㼐㼕㼚㼓㻌䝠䝖ᢠయ ᆺ㻌チฟ㢪䠖䠍䠅㻌䛾㑇ఏᏊኚ ᶵ 䛾ไᚚ䛻䜘䜛ኚ 䝍䞁䝟䜽䛾స〇᪉ἲ 㻌㻌㻌チ䠐䠐䠔䠓䠌䠒䠔㻌䠎䠅㻌 B 䛾ᢠయ㑇ఏᏊኚ ᵝᘧ䛾 ᪉ἲ 㻌㻌㻌㛤䠎䠌䠌䠕䠌䠒䠌䠔䠑䠌䠏䠅㻌 ᡤᮃ䛾䝫䝸䝨䝥䝏䝗䜢䛩䜛ᢠయ 䛾〇㐀᪉ἲ 㻌㻌㻌㢪䠎䠌䠌䠕䠎䠒䠐䠏98, US6/263,285 䠐䠅㻌

ELISA䛻䜘䜛ᢠཎ䜈䛾䛾 ᐃ ᯝ ㄽ 䠖 ) Kanayama, N. et al. Reversible switching of immunoglobulin hypermutation machinery in a chicken B cell line. Biochem. Biophys. Res. Commn. 327, 7-75 (25). 2) Kanayama, N.

Novel in vitro screening system for monoclonal antibodies using hypermutating chicken B cell library. J. Biosci. Bioeng. 2, 78-8 (26). ) Magari, M. et al.

10 ኚ ຊ䜢䛧䛯ᇵ㣴䠞䜢䛔䛯᪂つ䛺ᢠయぶ ᛶᡂ 䝅䝇䝔䝮㻰㼀㻠㻜㻙㻿㼃䛻䜘䜛 ᢠయ䛾ᶵ ᨵኚ㻔䝙䝽䝖䝸ᢠయᆺ㻕㻌㼂㻴㑇ఏᏊ㻌㑇ఏᏊ༢㞳㼂㻴㑇ఏᏊ ᛶ䜔ぶ ᛶ䛾 ᨵⰋ䛥䜜䛯mAb 㼂㻸㑇ఏᏊ㻌㼂㻸㑇ఏᏊ ᶵ ᨵኚ䛧䛯䛔 mab(䝠䝖䚸䝬䜴䝇) 䐟ᢠయ㑇ఏᏊᗙ䜈䛾㒊㑇ఏᏊᑟධ ኚ㒊 ᐃᖖ㒊 ᢠయ㑇ఏᏊᗙ እ ኚ㒊 እ᮶ᢠయ ኚ㒊䝍䞊䝀䝔䜱䞁䜾䝧䜽䝍䞊䝬䞊䜹䞊 loxp 䛘 ኚ ON 䜻䜻䝯䝷ᢠయ㻌䜻䝯䝷ᢠయ ᆺ㻌 ᆺ 㻰㼀㻠㻜㻙㻿㼃㻌㻰㼀㻠㻜㻙㻿㼃 Cre ぶぶ ᛶᡂ 䠄ኚ 䛸㑅ᢥ䠅䐠䝬䜴䝇mAb ኚ㒊䜢䛩䜛DT-SW 䝬䜴䝇ᢠ㻺㻼㼙㻭㼎䝬䜴䝇ᢠ㻺㻼㻌㼙㻭㼎㻌㻰㼀㻠㻜㻙㻿㼃㻌㻰㼀㻠㻜㻙㻿㼃㼙㻭㼎䝬䜴䝇ᢠ㻺㻼㻌㼙㻭㼎㻰㼀㻠㻜㻙㻿㼃㻌䝬䜴䝇ᢠ㻺㻼㻰㼀㻠㻜㻙㻿㼃 ᆺ㻌 ᆺ ኚ ON ~6 ᇵ㣴 5 2 FL-H 3 sigm FL2-H FL2-H % <APC-A> 5 NP binding <PE-A> 3 䝬䜴䝇VH 2 FL-H 3 Sorting 䐡DT-SW䛻䜘䜛䝬䜴䝇mAb䛾ぶ ᛶᡂ 䐡 DT-SW DT SW䛻䛻䜘䜛䝬䜴䝇mAb mab䛾䛾ぶ ᛶᡂ ぶ ᛶ䛜 Relative affinity y 㒊㒊㑇ఏᏊᑟධ <APC-A> Sorting 2 FL-H 3 2 FL-H Single clones Single clones ELISA䛻䜘䜛 ᑐぶ ᛶ䛾 Origin nal Original 3 3.% <PE-A> FL2-H FL2-H 䝬䜴䝇Vλ ᐃ ᯝ 䝠䝖ᆺ㻰㼀㻠㻜㻙㻿㼃䝅䝇䝔䝮㻌㻯㻴㻦䝠䝖㻯γ 㻝㻝㻌㻯㻴㻌㻦㻌䝠䝖㻯γ 㻯㻸㻦䝠䝖㻯κ 㻌㻯㻸㻌㻦㻌䝠䝖㻯κ 䜈䛾㻌䜈䛾㻰㼀㻠㻜㻙㻿㼃㻌㻰㼀㻠㻜㻙㻿㼃㻰㼀㻠㻜㻙㻿㼃㻙㼔㼓㻌㻰㼀㻠㻜㻙㻿㼃㻙㼔㼓㼓 ኚ 㻻㻺㻌 ኚ 㻻㻺㼂㻴㻘㼂㻸䛾㻌㻌㻜㻚㻡㻜㻚㻠䝠䝖IgGᆺᢠయ䝷䜲䝤䝷䝸䞊䛛䜙 ᢠཎ 䜽䝻䞊䞁䜢 ᚓ䛷䛝䜛㻜㻚㻟㻜㻚㻞㻜㻚㻝䝠䝖㻵㼓㻳㻝ᢠయ䛸䛧䛶ᶵ ᨵኚ 䝠䝖㻵㼓㻳㻝ᢠయ䛸䛧䛶ᶵ ᨵኚ㻌䠘䝠䝖ᆺᢠయ䛸䛧䛶 άᛶ䛾ホ౯䛜䠚 ᮏ 䛾 ᡂᯝ䛿䚸ᒸᒣTLO䜢䛨䛶( )ච ᕤᏛ ᡤ (௦ 䠖㻌 )䛻ᢏ 䛧䛶䛚䜚䚸ᐇ 䛻䛡䛶ඹ 㛤䜢㐍䜑䛶䛔䜛䚹䝠䝖IgG䜻䝯䝷ᢠయ䜢䐠䝠䝖IgGᆺᢠయ䝷䜲䝤䝷䝸䞊䛛䜙䛾ᢠཎ 䜽䝻䞊䞁䛾䝇䜽䝸䞊䝙䞁䜾㻺㻼㼎㼕㼚㼐㼕㼚㼓㻺㻼㻌㼎㼕㼚㼐㼕㼚㼓㻌䝠䝖ᢠయ ᆺ㻌チฟ㢪䠖䠍䠅㻌䛾㑇ఏᏊኚ ᶵ 䛾ไᚚ䛻䜘䜛ኚ 䝍䞁䝟䜽䛾స〇᪉ἲ 㻌㻌㻌チ䠐䠐䠔䠓䠌䠒䠔㻌䠎䠅㻌 B 䛾ᢠయ㑇ఏᏊኚ ᵝᘧ䛾 ᪉ἲ 㻌㻌㻌㛤䠎䠌䠌䠕䠌䠒䠌䠔䠑䠌䠏䠅㻌 ᡤᮃ䛾䝫䝸䝨䝥䝏䝗䜢䛩䜛ᢠయ 䛾〇㐀᪉ἲ 㻌㻌㻌㢪䠎䠌䠌䠕䠎䠒䠐䠏98, US6/263,285 䠐䠅㻌䝠䝖ᆺᢠయ䜢 ฟ䛩䜛䠞䛾స〇᪉ἲ 㻌㻌㻌㢪䠎䠌䠍䠌䠎䠑䠔䠐䠌䠐㻰㼀㻠㻜㻙㻿㼃㻌㻌㻰㼀㻠㻜㻙㻿㼃㻙㼔㼓㻌䝠䝖CJ (H㙐) 䝠䝖ᢠయ㻌 ᆺ㻌 ኚ 㻻㻺㻌䝠䝖㻵㼓㻳㻝ᆺᢠయ䝷䜲䝤䝷䝸䞊䝠䝖㻵㼓㻳㻝ᆺᢠయ䝷䜲䝤䝷䝸䞊㻌䛛䜙䛾ᢠయ᥈ 㻌䛛䜙䛾ᢠయ᥈ 䐟䝠䝖IgG䜻䝯䝷ᢠయ ᆺDT-SW 䝠䝖CN (L㙐) ᶵ ᨵኚ䛧䛯䛔mAb (䝠䝖䚸䝬䜴䝇) 䝠䝖䜻䝯䝷ᢠ య ᆺ 㻜 ELISA䛻䜘䜛ᢠཎ䜈䛾䛾 ᐃ ᯝ ㄽ 䠖 ) Kanayama, N. et al. Reversible switching of immunoglobulin hypermutation machinery in a chicken B cell line. Biochem. Biophys. Res. Commn. 327, 7-75 (25). 2) Kanayama, N. et al. Genetic manipulation of an exogenous non-immunoglobulin protein by gene conversion machinery in a chicken B cell line. Nucleic Acids Res. 3, e (26). 3) Todo, K. et al. Novel in vitro screening system for monoclonal antibodies using hypermutating chicken B cell library. J. Biosci. Bioeng. 2, 78-8 (26). ) Magari, M. et al. Enhancement of antibody production from a chicken B cell line DT by reducing Pax5 expression. J. Biosci. Bioeng. 7, (29). 5) Kajita, M. et al. Efficient affinity maturation of antibodies in an engineered chicken B cell line DT-SW by increasing point mutation. J. Biosci. Bioeng., (2). 6) Magari, M. et al. Enhancement of hypermutation frequency in the chicken B cell line DT for efficient diversification of the antibody repertoire. Biochem. Biophys. Res. Commn. 396, (2). ㄝ䠖㻌 ) 㻌㻌䚸㻌䛂ኚ ᶵ 䛾ON/OFFไᚚ 䛺B 䜢䛔䛯ᢠయ䛚䜘䜃ኚ 䝍䞁䝟䜽䛾స〇䝅䝇䝔䝮䛃㻌 ᐇ㦂 Ꮫ㻌㻌 Vol.2. p (26). 2) 㔠ᒣ ᶞ㻌䚸䛂ᇵ㣴B 䜢䛔䜛in vitroᢠయస〇䝅䝇䝔䝮䛻䜘䜛 ᢠయ䛾〇䛃㻌 YAKUGAKU ZASSHI 29, -7 (29). 䛚ၥ䛔䜟䛫ඛ䠖㻌 ᒸᒣ Ꮫ Ꮫ㝔 Ꮫ 㻌㔠ᒣ㻌 ᶞ TEL: nkanayam@cc.okayama-u.ac.jp

11 Lactobacillus plantarum 32 MIC DNA /2 * ( ) /6 ( ) 2 / ( ) 2/ ( ) 3 / ( ) / ( ) /5 ( 5 ) /2 ( ) 5 3/5 ( ) /7 ( ) DNA /6 ( ) 2/6 ( 3 ) kb Ladder Lactobacillus plantarum Kitek Traditional fermented milk in Xinjiang Uiguru autonomy, China. One of the characteristics of this fermented milk is its viscosity Phylogeny of wild Lactobacillus strain 975 as determined by 6S rdna sequence analysis Escherichia coli Penicillium olsonii Lactobacillus vaginalis Lactobacillus delbrueckii ssp. bulgaricus Lactobacillus delbrueckii ssp. lactis Strain 975 Lactobacillus delbrueckii ssp. delbrueckii Lactobacillus ensenii Lactobacillus gasseri Lactobacillus acetotolerans Lactobacillus helveticus Lactobacillus acidophilus Lactobacillus amylovorus

12 (3~5) ~2(2~25) () Hurunge ACE κ- QEQNQEQPIRCEKDERFFSDKIAKYIPIQYVLSRYPSYGLNYYQQKPVAL 5 INNQFLPYPYYAKPAAVRSPAQILQWQVLSNTVPAKSCQAQPTTMARHPH PHLSFM AIPPKKNQD KTEIPTINTIASGEPTSTPTIEAVESTVATLEASP Ala-Ile-Pro-Pro-Lys-Lys-Asn-Gln-Asp 5 EVIESPPEIN TVQVTSTAV 69 κ-κcn 7-5) Retention time (min) Lb. helveticus 3B RP-HPLCACE SE B MSC3 MSC7 MSC3 MSC8 DW well 5μ HPLC Column Shim-pack VP-ODS (5mmL..6mmI.D.) Mobile phase 2mM sodium phosphate buffer(ph2.5) containing mm sodium -hexanesulfonate Flow rate.8ml/min Temperature 5 Reaction reagent o-phthalaldehyde reagent Flow rate.ml/min Temperature 5 GABA V.cholerae MRSA A 2 B 2 E.coli IFO E.coli O57 L.m M2- Activity spectra of antibacterial substance produced by Lactococcus lactis subsp. lactis KM Indicator strain Inhibitory zone(mm) Lb.acidophilus NIAI L-5 6 Lb.helveticus 3M2 2 Lb.casei ATCC393 3 Lb.delbrueckii subsp.delbrueckii ATCC Lb.plantarum IFO37 8 Lb.amylovorus IMC- 22 Lc.garvieae 79(38) 9 Lc.lactis subsp.lactis KM Lc.lactis subsp.lactis Leuc.mesenteroides Sc.thermophilus 253B2 22 Ec.faecalis RIMD36 9 Ec.faecium C2 Pseudomonas fragi IFO358 Staphylococcus aureus IAM 2 Staphylococcus piscifermentans GT2-23 Bacillus subtilis IFO325 Listeria monocytogenes VTU26 7 Escherichia coli RB GABA2723 GABA 5.6mg/ml Lactococcus lactis subsp. lactis KM.5% Na% GABA.55mg/ml C 2 GABA.6mg/ml n-32.7% 6F25 :2:3:No.3 :5:6:7: 8:GABA(2.5mg/ml)9:Na(.6mg/ml) KMGABA TELFAX tmnono@cc.okayama-u.ac.jp

12 九州産業大学情報科学会誌 10 巻 1 号 (2011 年 11 月 43) 2011 Koichiro Tanaka Department of Information Science, Faculty of Information Science, Kyushu Sangyo Univer

12 九州産業大学情報科学会誌 10 巻 1 号 (2011 年 11 月 43) 2011 Koichiro Tanaka Department of Information Science, Faculty of Information Science, Kyushu Sangyo University tanaka@is.kyusan-u.ac.jp, http://www.is.kyusan-u.ac.jp/~tanaka/