簡易操作ガイド

Applied Biosystems
StepOne and StepOnePlus
Real Time PCR System 簡易操作ガイド
StepOne Software v2.3
2015 年 7 月 Rev. H

For Research Use Only. Not for use in diagnostic procedures.

᧯స➼䛾ヲ⣽䛿ᚲ䛪〇ရ䛻௜ᒓ䛾䝴䞊䝄䞊䜺䜲䝗䠄䝬䝙䝳䜰䝹䠅䜢ཧ↷䛟䛰䛥䛔䚹グ㏙ෆᐜ䛿ᨵⰋ䛾䛯䜑ண࿌䛺䛟ኚ᭦䛥䜜䜛ሙྜ䛜䛒䜚䜎䛩䛾䛷䚸䛤஢ᢎୗ䛥䛔䚹〇ရ䛾௙ᵝ䞉እほ䛿ᨵⰋ䛾䛯䜑ண࿌䛺䛧䛻ኚ᭦䛥䜜䜛ሙྜ䛜䛒䜚䜎䛩䚹

-1-

┠ḟ
䝽䞊䜽䝣䝻䞊

3

StepOne / StepOnePlus Real-Time PCR System 䛾㉳ື4Standard Curve Method ᪂つᐇ㦂⣔䛾సᡂ : Design Wizard 䜢⏝䛔䛯᪉ἲ

5Standard Curve Method ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ17཯ᛂ䝥䝺䞊䝖䛾‽ഛ

26

䝷䞁䛾㛤ጞ䛸⤊஢

29

Standard Curve Method 䝕䞊䝍䛾ゎᯒ䞉ฟຊ

34

Relative Standard Curve ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ

45

Relative Standard Curve 䝕䞊䝍䛾ゎᯒ

47

Comparative CT (ǻǻCT) ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ

48

Comparative CT (ǻǻCT) 䝕䞊䝍䛾ゎᯒ : ༢⊂䝗䜻䝳䝯䞁䝖ෆ䛷䛾ゎᯒ

51

Comparative CT (ǻǻCT) 䝕䞊䝍䛾ゎᯒ : Study 䜢⏝䛔䛯༢⊂ཪ䛿」ᩘᯛ䛾ゎᯒ

52

Appendix A 䠖 StepOnePlus System 䛷䛾 VeriFlex ᶵ⬟䛾タᐃ᪉ἲ

59

Appendix B 䠖 䝍䝑䝏䝟䝛䝹᧯స᪉ἲPCĺStepOne 䝅䝇䝔䝮䜈䛾䝣䜯䜲䝹䛾㌿㏦䛸䝷䞁

61ಖᏑ Template 䝣䜯䜲䝹䜢࿧䜃ฟ䛩᪉ἲ

65

Appendix C 䠖ᮏయ䛛䜙䝕䞊䝍䜢ᅇ཰䛩䜛᪉ἲ

Standard Curve
Relative
Standard Curve᳨㔞⥺䜢⏝䛔䛯ᐃ㔞᳨㔞⥺䜢⏝䛔䛯
┦ᑐⓎ⌧ᐃ㔞

Comparative
CT(ǻǻCT)䝃 䜲 䜽 䝹ẚ ㍑ἲ
䜢⏝䛔䛯┦ᑐⓎ⌧ᐃ㔞Comparative
CT(ǻǻCT) 㻌Study䝃 䜲 䜽 䝹ẚ ㍑ἲ
䜢⏝䛔䛯┦ᑐⓎ⌧ᐃ㔞63᳨㔞⥺䜢సᡂ䛧䚸䛭䜜䛮䜜䛾᳨య䛾ᐃ㔞⤖ᯝ䜢ᑟ䛝ฟ䛩᪉ἲ䛷䛩䚹⤯ᑐᐃ㔞䜢⾜䛔䛯䛔ሙྜ䛿䛣䛾᪉ἲ䜢⏝䛔䜎䛩䚹ᶆⓗ㑇ఏᏊ,ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ䛾᳨㔞⥺䛸᳨య䛜ྠ୍䝥䝺䞊䝖ෆ䛻Ꮡᅾ䛩䜛ሙྜ䚸ྛ᳨య䛾ྛ㑇ఏᏊ䛾ᐃ㔞⤖ᯝ䜢⟬ฟ䛧䛯ᚋ䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ䛷ᶆ‽໬䛥䜜䛯ᶆⓗ㑇ఏᏊ䛾┦ᑐⓎ⌧ẚ䜢⟬ฟ䛧䜾䝷䝣໬䛩䜛ゎᯒ᪉ἲ䛷䛩䚹ᐃ䛧䛯䛔᳨య䛾ᶆⓗ㑇ఏᏊ䛸ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ䛜ྠ୍䝥䝺䞊䝖ෆ䛻Ꮡᅾ䛩䜛ሙྜ䚸᳨ฟ䝃䜲䜽䝹(CT ್)䛾ᕪ䜢⏝䛔䛶ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ䛷ᶆ‽໬䛥䜜䛯ᶆⓗ㑇ఏᏊ䛾┦ᑐⓎ⌧ẚ䜢⟬ฟ䛧䜾䝷䝣໬䛩䜛ゎᯒ᪉ἲ䛷䛩䚹Comparative CT (ǻǻCT)䛾䝥䝺䞊䝖䜢 1 ᯛ䡚」ᩘᯛྠ᫬䛻ゎᯒ䛩䜛᪉ἲ䛷䛩䚹䛣䛾᪉ἲ䛷䛿 Biological Replicate Group 䜢ᣦᐃ䛧䛶ゎᯒ䛩䜛䛣䛸䛜ྍ⬟䛷䛩䠄49 䝨䞊䝆ཧ↷䠅䚹

-2-

JP-886̲本文̲0707se.indd 1

15/07/08 17:33

䝽䞊䜽䝣䝻䞊
Applied Biosystems™ StepOne™ and StepOnePlus™ Real-Time PCR System 䜢⏝䛔䛶ᐇ㦂䜢⾜䛖䛸䛝䛾䝽䞊䜽䝣䝻䞊䛿௨ୗ䛾㏻䜚䛷䛩䚹㟁※䜢ධ䜜䜛䛸⮬ືⓗPC 䛾㟁※䜢ධ䜜䜛䛻䝅䝇䝔䝮デ᩿䜢⾜䛔䜎䛩䚹⣙ 10 ศせ䛧䜎䛩

StepOne 䜎䛯䛿 StepOnePlus 䛾㟁※䜢ධ䜜䜛

StepOne 䝋䝣䝖䜴䜵䜰䜢㉳ື䝗䜻䝳䝯䞁䝖䛾タᐃ䛿䝷䞁୰䜔䝷᪂つᐇ㦂⣔䛾సᡂ䞁⤊஢ᚋ䛻⾜䛖䛣䛸䜒䛷䛝䜎䛩཯ᛂ⣔䛾ㄪᩚ / ཯ᛂ䝥䝺䞊䝖䛾‽ഛ

䝷䞁䛾㛤ጞ

䝷䞁䛾⤊஢

䝕䞊䝍ゎᯒ䞉ฟຊ

StepOne 䝋䝣䝖䜴䜵䜰䜢⤊஢PC 䛾㟁※䜢ษ䜛-3-

JP-886̲本文̲0707se.indd 2

15/07/08 17:33

StepOne / StepOnePlus Real-Time PCR System 䛾㉳ື1.

䝁䞁䝢䝳䞊䝍䛾㟁※䜢ධ䜜䜎䛩䚹

2.

StepOne 䜎䛯䛿 StepOnePlus ᶵჾ䛾㟁※䜢ධ䜜䜎䛩䚹ᮏయ⫼㠃䛻䝇䜲䝑䝏䛜䛒䜚䜎䛩䚹

㻌㻌㻌㻌㻌
㻌㻌
ὀグ䠖㟁※䜢ධ䜜䜛䛸⮬ືⓗ䛻䝅䝇䝔䝮デ᩿䜢⾜䛔䜎䛩䠄⣙ 10 ศ䠅䚹ᮏయ䛿᭱ᚋ䛾᧯సᚋ 4 ᫬㛫䠄ึᮇタᐃ䠅⤒㐣䛩䜛䛸⮬ືⓗ䛻䝇䝸䞊䝥䝰䞊䝗䛻ධ䜚䜎䛩䚹⏬㠃ᕥୗ䛾䝪䝍䞁䜢䝍䝑䝏䛩䜛䛸䛩䛠䛻䝇䝸䞊䝥䝰䞊䝗䛻䛷䛝䜎䛩䚹䝇䝸䞊䝥䝰䞊䝗䜢ゎ㝖䛩䜛䛸䛝䜒䛣䛾䝪䝍䞁䜢䝍䝑䝏䛧䜎䛩䚹⿵㊊䠖ゎᯒ⏝䛾䝁䞁䝢䝳䞊䝍䛻䝋䝣䝖䜴䜵䜰䜢䜲䞁䝇䝖䞊䝹䛧䛯ሙྜ

StepOne 䝋䝣䝖䜴䜵䜰䛿ึᅇ㉳ື᫬䛻 StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛸䛾䝁䝭䝳䝙䜿䞊䝅䝵䞁䛾☜ㄆ䜢⾜䛔䜎䛩䚹ゎᯒ⏝䛾䝁䞁䝢䝳䞊䝍䛷䛿ᚲせ䛺䛔సᴗ䛺䛾䛷䚸ୗグ䛾ᡭ㡰䛷䛣䛾タᐃ䜢ゎ㝖䛧䛶䛟䛰䛥䛔䚹ⱝᖸ㉳ື䝇䝢䞊䝗䛜㏿䛟䛺䜚䜎䛩䚹
1.

StepOne 䝋䝣䝖䜴䜵䜰䜢㉳ື䛧䜎䛩䚹

2.

Login ⏬㠃䛜⾲♧䛥䜜䛯䜙䚸Guest ௨እ䛾 User Name 䜢㑅ᢥ䛧䚸OK 䜢䜽䝸䝑䜽䛧䜎䛩䚹ὀグ䠖Guest 䛷䝻䜾䜲䞁䛩䜛䛸 4 䛾䝏䜵䝑䜽䝪䝑䜽䝇䛜䜰䜽䝔䜱䝤䛻䛺䜙䛪䚸タᐃኚ᭦䛩䜛䛣䛸䛜䛷䛝䜎䛫䜣䚹3.

Tools > Preferences 䜢㑅ᢥ䛧䚸Startup 䝍䝤䜢䜽䝸䝑䜽䛧䜎䛩䚹4.

Connect to Default instrument 䛾䝏䜵䝑䜽䝪䝑䜽䝇䜢䛿䛪䛧䜎䛩䚹

5.

OK 䜢䜽䝸䝑䜽䛧䜎䛩䚹-4-

JP-886̲本文̲0707se.indd 3

15/07/08 17:33

Standard Curve Method ᪂つᐇ㦂⣔䛾సᡂ : Design Wizard 䜢⏝䛔䛯᪉ἲ
StepOne 䝋䝣䝖䜴䜵䜰䛾 Design Wizard 䜢⏝䛔䛶᪂つᐇ㦂⣔䜢సᡂ䛧䜎䛩䚹

1.

StepOne 䝋䝣䝖䜴䜵䜰䛾䝅䝵䞊䝖䜹䝑䝖䜢䝎䝤䝹䜽䝸䝑䜽䛧䛶䚸䝋䝣䝖䜴䜵䜰䜢㉳ື䛧䜎䛩䚹ὀグ䠖StepOne 䝋䝣䝖䜴䜵䜰䛾㉳ື䛻䛿⣙ 15 ⛊⛬ᗘ䛛䛛䜚䜎䛩䚹

2.

Login ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹୍᫬ⓗ䛻䝻䜾䜲䞁䛩䜛䛸䛝䛻䛿 Log in as Guest 䜢䜽䝸䝑䜽䛧䜎䛩䚹䝴䞊䝄䞊Ⓩ㘓䛧
䛶౑⏝䛩䜛ሙྜ䛿䚸᪂䛧䛔 User Name 䜢䝍䜲䝥ධຊ䛧䚸OK 䜢䜽䝸䝑䜽䛧䜎䛩䚹㻌ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹ὀグ䠖୍ᗘⓏ㘓䛧䛯䝴䞊䝄䞊ྡ䛿䚸ḟᅇ䛛䜙䝥䝹䝎䜴䞁䝯䝙䝳䞊䜘䜚㑅ᢥ䛷䛝䜛䜘䛖䛻䛺䜚䜎䛩䚹3.

䝩䞊䝮⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
Design Wizard 䜢䜽䝸䝑䜽䛧䜎䛩䚹-5-

JP-886̲本文̲0707se.indd 4

15/07/08 17:33

4.

1A. Define Experiment Properties ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹ྛ㡯┠䛾ධຊ䞉㑅ᢥ䜢⾜䛔䜎䛩䚹- Experiment Name 䛾ධຊ䛿ᚲ㡲䛷䛩䚹䛣䛾ྡ๓䛜䛭䛾䜎䜎䝣䜯䜲䝹ྡ䛻䛺䜚䜎䛩䚹- 䛭䛾䜋䛛䛾㡯┠䛾ධຊ䛿௵ព䛷䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹- Which instrument are you using to run the experiment 䛷䛿䚸StepOne(48well) 䜎䛯䛿
StepOnePlus(96well) 䛾䛹䛱䜙䛛䚸䛤౑⏝䛾ᶵჾ䜢㑅ᢥ䛧䜎䛩䚹㻌 ὀグ䠖㛫㐪䛳䛯ᶵჾ䜢㑅ᢥ䛧䛯ሙྜ཯ᛂ䜢ጞ䜑䜛䛣䛸䛜䛷䛝䜎䛫䜣䚹- What Type of Experiment do you want to design 䛾㡯┠䛿䚸ᐃ㔞䜰䝑䝉䜲䜢⾜䛖䛸䛝䛻䛿 Quantitation 䜢㑅ᢥ䛧䜎䛩䚹タᐃ䛜⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹-6-

JP-886̲本文̲0707se.indd 5

15/07/08 17:33

5.

1B. Define the Methods and Materials ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹ྛ㡯┠䛾㑅ᢥ䜢⾜䛔䜎䛩䚹ὀグ䠖Run Method ௨እ䛾඲䛶䛾タᐃ䛿䝷䞁୰ཬ䜃䝷䞁⤊஢ᚋ䛷䜒ኚ᭦䛷䛝䜎䛩䚹- quantitation method :㻌 ᐃ㔞䜢䛹䛾ᡭἲ䛷⾜䛖䛛㑅ᢥ䛧䜎䛩䚹㻌 Standard Curve : ཯ᛂ⣔ẖ䛻᳨㔞⥺䜢సᡂ䛧䚸ᐃ㔞್䜢⟬ฟ䛧䜎䛩
㻌 Relative Standard Curve : ཯ᛂ⣔ẖ䛻᳨㔞⥺䛛䜙ᐃ㔞್䜢ồ䜑䛯䛒䛸䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛾್䛷䛾⿵ṇཬ䜃䜻䝱䝸䝤䝺䞊䝍䝃䞁䝥䝹䛸䛾┦ᑐ್䜢䚸䝋䝣䝖䜴䜵䜰䛜⮬ືⓗ䛻⟬ฟ䛧䜎䛩䚹
㻌 Comparative CT (ǻǻCT) : ẚ㍑ CT ἲ䛷䛾┦ᑐᐃ㔞䜢䝋䝣䝖䜴䜵䜰䛜⮬ືⓗ䛻⟬ฟ䛧䜎䛩䚹- reagents : ཯ᛂ⣔䛾䜿䝭䝇䝖䝸䜢㑅ᢥ䛧䜎䛩䚹㻌 SYBR Green Reagents 䜢㑅ᢥ䛩䜛䛸ᕥୗ䛻 Include Melt Curve 䛾䝏䜵䝑䜽䝪䝑䜽䝇䛜⾲♧䛥䜜䜛䛾䛷䚸㻌 䝏䜵䝑䜽䛜ධ䛳䛶䛔䜛䛣䛸䜢䛤☜ㄆ䛟䛰䛥䛔䚹- ramp speed : Standard 䛸 Fast 䛛䜙㑅ᢥ䛧䜎䛩䚹㻌 䛤฼⏝䛻䛺䜛཯ᛂヨ⸆䛻ྜ䜟䛫䛶㑅ᢥ䛧䜎䛩- template : ౑⏝䛩䜛䝃䞁䝥䝹䛾✀㢮䜢㑅ᢥ䛧䜎䛩䚹඲䛶䛾タᐃ䛜⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹-7-

JP-886̲本文̲0707se.indd 6

15/07/08 17:33

6.

2A. Set Up: Targets ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹6a䝍䞊䝀䝑䝖䠄䝥䝷䜲䝬䞊䝉䝑䝖䠅䛾ᩘ䜢ධຊ䛧䜎䛩䚹6b᳨㔞⥺䜢タᐃ䛩䜛᫬䛻䛿䝏䜵䝑䜽䜢ධ䜜䜎䛩䚹6c┠ⓗ㑇ఏᏊ䛾ྡ๓䜢ධຊ䛧䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹6d

TaqMan™ 䝥䝻䞊䝤䛾䝸䝫䞊䝍䞊⺯ගⰍ⣲䜎䛯䛿 SYBR™䛾䛔䛪䜜䛛䜢䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛧䜎䛩䚹6e䜽䜶䞁䝏䝱䞊䛾ධຊ㡯┠䛷䛩䚹TaqMan 䝥䝻䞊䝤䛾ሙྜ䛿 TAMRA™䚸TaqMan MGB 䝥䝻䞊䝤䛾ሙྜ䛿 NFQ-MGB䚸SYBR Green 䜰䝑䝉䜲䛾ሙྜ䛿 None 䜢タᐃ䛧䜎䛩䚹ὀグ䠖StepOne 䝅䝇䝔䝮䛷䛿䚸TAMRA Ⰽ⣲䜢⏝䛔䛯ᐇ㦂䜢⾜䛖䛣䛸䛿䛷䛝䜎䛫䜣䚹

6f

䝍䞊䝀䝑䝖ẖ䛾⾲♧Ⰽ䜢㑅ᢥ䛧䜎䛩䚹

6a

6b

6c

6d

6e

6f

タᐃ䛜⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹-8-

JP-886̲本文̲0707se.indd 7

15/07/08 17:33

7.

2B. Set Up: Standard ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹7a᳨㔞⥺సᡂ䛾䛯䜑䛾ᕼ㔘䝃䞁䝥䝹䛾ᩘ䜢ධຊ䛧䜎䛩䚹ὀグ䠖䜰䝥䝷䜲䝗䝞䜲䜸䝅䝇䝔䝮䝈䛷䛿 5 Ⅼタᐃ䛩䜛䛣䛸䜢᥎ዡ䛧䜎䛩䚹7bྠ୍䝃䞁䝥䝹䛾཯᚟ᩘ䜢ධຊ䛧䜎䛩䚹7c᳨㔞⥺సᡂ䝃䞁䝥䝹䛾᭱኱್䜎䛯䛿᭱ᑠ್䜢ධຊ䛧䜎䛩䚹7dᕼ㔘ಸ⋡䜢䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛧䜎䛩䚹- 1:5 䜢㑅ᢥ䛩䜛䛸䚸㉳Ⅼ䛾⃰ᗘ䛻ᑐ䛧䛶 1/5 䛾್䛜⧞䜚㏉䛧⮬ືⓗ䛻⟬ฟ䛥䜜䜎䛩䚹- X10 䜢㑅ᢥ䛩䜛䛸䚸㉳Ⅼ䛾⃰ᗘ䛻ᑐ䛧䛶 10 ಸ䛾್䛜⧞䜚㏉䛧⮬ືⓗ䛻⟬ฟ䛥䜜䜎䛩䚹

7eStandard Curve Preview 䜢☜ㄆ䛧䜎䛩䚹

7f

䝃䞁䝥䝹䜢䛹䛾୪䜃䛻タᐃ䛩䜛䛛䛾᪉ྥ䜢タᐃ䛧䜎䛩䠄ึᮇタᐃ䛿ᶓ᪉ྥ䠅䚹

7g

Negative Control 䛾タᐃ఩⨨䜢䚸ᕥୖ䛸ྑୗ䛛䜙㑅ᢥ䛷䛝䜎䛩䠄ึᮇタᐃ䛿ᕥୖ䠅䚹
㻌

7g

7f

7a
7b

7c

7d

7e

㻌

㻌㻌
඲䛶☜ㄆ䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹㻌 㻌-9-

JP-886̲本文̲0707se.indd 8

15/07/08 17:33

8.

2C. Set Up: Samples ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹8aᮍ▱䝃䞁䝥䝹䛾ᩘ䜢ධຊ䛧䜎䛩䚹8bྠ୍䝃䞁䝥䝹䛾཯᚟ᩘ䜢ධຊ䛧䜎䛩䚹

8c

Negative Control 䛾䜴䜵䝹䛾ᩘ䜢ධຊ䛧䜎䛩䚹8d8a 䛷ධຊ䛧䛯䝃䞁䝥䝹ᩘ䛻ᛂ䛨䛶䝸䝇䝖䛜⾲♧䛥䜜䜛䛾䛷䚸䝃䞁䝥䝹ྡ䜢ධຊ䛧䚸Color 䜢タᐃ䛧䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹8e඲䛶䛾䝃䞁䝥䝹䜢඲䛶䛾䝍䞊䝀䝑䝖䛻ᑐ䛧䛶཯ᛂ䛩䜛䛛䚸䜎䛯䛿䝃䞁䝥䝹ẖ䛻䝍䞊䝀䝑䝖䜢タᐃ䛩䜛䛛䚸㑅ᢥ䛧䜎䛩䚹

8f

ୖグධຊ䛜඲䛶⤊஢䛩䜛䛸 Unknown䚸Standard䚸Negative Control ཬ䜃✵䛝䜴䜵䝹䛾ᩘ䛜⮬ືⓗ䛻⟬ฟ䛥䜜䚸⾲♧䛥䜜䜎䛩䚹8g

䝃䞁䝥䝹䜢䛹䛾୪䜃䛻タᐃ䛩䜛䛛䛾᪉ྥ䜢タᐃ䛧䜎䛩䠄ึᮇタᐃ䛿ᶓ᪉ྥ䠅䚹

8h

Negative Control 䛾タᐃ఩⨨䜢䚸ᕥୖ䛸ྑୗ䛛䜙㑅ᢥ䛷䛝䜎䛩䠄ึᮇタᐃ䛿ᕥୖ䠅䚹

8g

8f

8a
8b
8c

8d

8e

8f

඲䛶䛾タᐃ䛜⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹- 10 -

JP-886̲本文̲0707se.indd 9

15/07/08 17:33

9.

2D. Set Up: Run Method ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
9aGraphical View 䝍䝤䛛 Tabular View 䝍䝤䜢㑅ᢥ䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Graphical View 䝍䝤⏬㠃䜢⏝䛔䛶ㄝ᫂䛧䜎䛩䚹9b཯ᛂ䝪䝸䝳䞊䝮䜢ධຊ䛧䜎䛩䚹StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛷䛿 10-30ȝL 䛾⠊ᅖෆ䛷ᐇ㦂䜢⾜䛖䛣䛸䜢᥎ዡ䛧䛶䛔䜎䛩䚹ὀグ䠖ᐇ㦂ྍ⬟䛺᭱኱䝪䝸䝳䞊䝮䛿 50μL 䛷䛩䚹31μL ௨ୖ䛻タᐃ䛩䜛䛸䝷䞁䝥䝇䝢䞊䝗䛜㐜䛟䛺䜚䜎䛩䚹9cᗘ᮲௳䜢☜ㄆ䛧䜎䛩䚹ᚲせ䛻ᛂ䛨䛶䚸ᕥୖ䛾䝪䝍䞁䜢㑅ᢥ䛩䜛䛣䛸䛷䚸௵ព䛾䝥䝻䜾䝷䝮䜢㏣ຍ䞉๐㝖䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹9d䛾䝬䞊䜽䛜䜰䜽䝔䜱䝤䛻䛺䛳䛶䛔䜛䝫䜲䞁䝖䛷 StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛿⺯ග䝕䞊䝍䜢ᅇ཰䛧䜎䛩䚹䝬䞊䜽䛾ୖ䜢䜽䝸䝑䜽䛩䜛䛣䛸䛷䚸䝕䞊䝍ᅇ཰䝫䜲䞁䝖䜢タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹ὀグ䠖䝃䜲䜽䝹䝇䝔䞊䝆ෆ䛷䛾䝕䞊䝍ᅇ཰䝫䜲䞁䝖䛿ᚲ䛪 1 䞄ᡤ䛻䛺䜛䜘䛖䛻䛤ὀព䛟䛰䛥䛔䚹ὀグ䠖䝕䞊䝍ᅇ཰䝫䜲䞁䝖䛾䝬䞊䜽䛜䚸䝃䜲䜽䝹䝇䝔䞊䝆䛾᭱ᚋ䛾䝇䝔䝑䝥䛷䜰䜽䝔䜱䝤䛻䛺䛳䛶䛔䜛䛣䛸䜢䛤☜ㄆ䛟䛰䛥䛔䚹䛣䛾䝬䞊䜽䛿 1 ᅇ䜽䝸䝑䜽䛩䜛䛸㠀䜰䜽䝔䜱䝤䛻䛺䜚䚸䛭䛾䜎䜎䝷䞁䜢⾜䛖䛸䝕䞊䝍ᅇ཰䛜⾜䜟䜜䜎䛫䜣䚹

9e

Run Method 䛾᮲௳䜢ಖᏑ䛩䜛ሙྜ䛻䛿 Save Run Method 䜢䜽䝸䝑䜽䛧䜎䛩䚹ಖᏑ䛧䛯 Run Method䛿䚸ḟᅇ௨㝆ྠ䛨᮲௳䛷ᐇ㦂䜢⾜䛖䛸䛝䛻 Open Run Method 䜢䜽䝸䝑䜽䛧䛶࿧䜃ฟ䛩䛣䛸䛜䛷䛝䜎䛩䚹

9a

9b

9e
9c

9d

඲䛶䛾タᐃ䛜⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹- 11 -

JP-886̲本文̲0707se.indd 10

15/07/08 17:33

10. 2E. Set Up: Reaction Setup > Reaction Mix Calculations ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
10a Reaction Mix Calculations 䜢㑅ᢥ䛧䜎䛩䠄ึᮇタᐃ䠅䚹
10b ┠ⓗ㑇ఏᏊ䛾཯ᛂ⣔䜢㑅ᢥ䛧䜎䛩䚹
10c 䝥䝷䜲䝬䞊䞉䝥䝻䞊䝤䛾ヨ⸆䛾ᙧែ䛻ᛂ䛨䛶 Assay Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛧䜎䛩䚹Inventoried / Made to Order : TaqMan Gene Expression Assays 䜎䛯䛿ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ⏝ヨ⸆䛷䚸䝥䝷䜲䝬䞊䞉䝥䝻䞊䝤䛜 20X 䛾 Mixture 䛻䛺䛳䛶䛔䜛ヨ⸆䜢䛤฼⏝䛾ሙྜ
Custom : 䜹䝇䝍䝮䛷タィ䞉ྜᡂ䜢⾜䛔䚸䝥䝷䜲䝬䞊䞉䝥䝻䞊䝤䛜ู䚻䛾䝏䝳䞊䝤䛻ධ䛳䛶䛔䜛ሙྜ
10d 1 䜴䜵䝹䛒䛯䜚䛾཯ᛂ䝪䝸䝳䞊䝮䜢ධຊ䛧䜎䛩䚹
10e 䝢䝨䝑䝔䜱䞁䜾䝻䝇ศ䜢⪃៖䛧䛯䝥䝷䝇 Į ศ䜢䛹䛾⛬ᗘ䛻䛩䜛䛛タᐃ䛧䜎䛩䚹
10f PCR ཯ᛂヨ⸆䠄Master Mix 䝍䜲䝥䛻䛾䜏ᑐᛂ䠅䛾⃰ᗘ䜢ධຊ䛧䜎䛩䚹10g 䝥䝷䜲䝬䞊䞉䝥䝻䞊䝤䛾⃰ᗘ䜢ධຊ䛧䜎䛩䚹10c 䛷 Custom 䜢㑅ᢥ䛧䛯ሙྜ䛿䚸䝥䝷䜲䝬䞊ཎᾮ䛾⃰ᗘ䜢䚸཯ᛂᾮ୰䛾⤊⃰ᗘ䜢ධຊ䛧䜎䛩䚹
10h ⮬ືⓗ䛻 1 䜴䜵䝹䛒䛯䜚䛾ヨ⸆䛾ㄪᩚ㔞䛜⾲♧䛥䜜䜎䛩䚹Print Reaction Setup 䜢䜽䝸䝑䜽䛧 Pre View䜢㛤䛟䛸䚸᭱⤊ㄪᩚ㔞䜢☜ㄆ䛩䜛䛣䛸䛜ฟ᮶䜎䛩䚹䝥䝸䞁䝖䜰䜴䝖䛻䛴䛔䛶䛿ḟ䝨䞊䝆 10o 䜢ཧ↷䛟䛰䛥䛔䚹
10i᳨㔞⥺సᡂ䝃䞁䝥䝹䛾ཎᾮ䛾⃰ᗘ䜢ධຊ䛧䜎䛩䚹10j༢఩䜢ධຊ䛧䜎䛩䚹ng 䛸 ȝg 䛿䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛷䛝䜎䛩䚹䛭䜜௨እ䛾༢఩䠄౛䛘䜀 copies䠅䛾᫬䛿┤᥋ධຊ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

10k ᳨㔞⥺䝃䞁䝥䝹䛾ᕼ㔘ẁ㝵䜢సᡂ䛩䜛㔞䛜⟬ฟ䛥䜜䜎䛩䚹

10a

10c

10d

10e
10h

10b
10g
10f

10h

10i

10j

10k

- 12 -

JP-886̲本文̲0707se.indd 11

15/07/08 17:33

10l

Sample Dilution Calculations 䝍䝤䜢䜽䝸䝑䜽䛧䜎䛩䚹

10m ཯ᛂ⣔䛻ຍ䛘䜛䝃䞁䝥䝹䛾⃰ᗘ䜢ධຊ䛧䜎䛩䚹10n 䝃䞁䝥䝹ẖ䛻ཎᾮ䛾⃰ᗘ䜢ධຊ䛩䜛䛸䚸10m 䛷タィ䛧䛯⃰ᗘ䛻ྜ䜟䛫䜛䛯䜑䛾ィ⟬䛜⮬ືⓗ䛻⾜䜟䜜
䜎䛩䚹
10m
10l

10o

10n

10o 䝥䝸䞁䝍䞊䛜᥋⥆䛥䜜䛶䛔䜛ሙྜ䚸タᐃ䛧䛯཯ᛂ⣔ㄪᩚ᮲௳䜢༳ๅ䛷䛝䜎䛩䚹༳ๅ䛩䜛䛸䛝䛻䛿䚸Print Reaction Setup 䜢䜽䝸䝑䜽䛧䜎䛩䚹
10p Print Reaction Setup Instructions ⏬㠃ෆ䛷䚸ୗグ㡯┠䜢㑅ᢥ䛧䜎䛩䚹- Detailed Reaction Setup Instructions- Include Plate Layout- Use sample color
10q Print 䜢䜽䝸䝑䜽䛧䜎䛩䚹䝥䝸䞁䝍䞊䛾☜ㄆ⏬㠃䛜⾲♧䛥䜜䜛䛾䛷 OK 䜢䜽䝸䝑䜽䛩䜛䛸༳ๅ䛥䜜䜎䛩䚹

10p

10q

඲䛶⤊஢䛧䛯䜙 Next>䜢䜽䝸䝑䜽䛧䜎䛩䚹- 13 -

JP-886̲本文̲0707se.indd 12

15/07/08 17:33

11. 3A. (Optional) Order: Materials List ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
11a ὀグ䠖䛣䛾సᴗ䜢⾜䛖䛯䜑䛻䛿䚸䛤฼⏝䛾 PC 䛜䜲䞁䝍䞊䝛䝑䝖䛻᥋⥆䛧䛶䛔䜛䛣䛸䛜ᚲせ䛷䛩䚹
Fine Assay 䛾 Enter Gene Name 䛻┠ⓗ㑇ఏᏊ䛾 Gene Name 䜔 Accession No.䜢ධຊ䛧 Find
Assay 䜢䜽䝸䝑䜽䛩䜛䛸䚸TaqMan Gene Expression Assay ෆ䛾䝥䝷䜲䝬䞊䞉䝥䝻䞊䝤䝉䝑䝖䛾᳨⣴䜢⾜䛔䚸䛒䜛ሙྜ䛻䛿䝸䝇䝖䛜⾲♧䛥䜜䜎䛩䚹
11b Experiment Materials List 䛻䛿䚸ᐇ㦂䜢⾜䛖䛯䜑䛻ᚲせ䛺ᾘ⪖ရ䛸䚸㑅ᢥ䛧䛯䜿䝭䝇䝖䝸䛻ᑐᛂ䛩䜛཯ᛂ
ヨ⸆䛜䝸䝇䝖䜰䝑䝥䛥䜜䜎䛩䚹11a 䛷 TaqMan Gene Expression Assay 䜢᳨⣴䞉㑅ᢥ䛧䛯ሙྜ䚸୍␒ୗẁ
䛻㏣ຍ䛷⾲♧䛥䜜䜎䛩䚹11c ᪥ᮏ䛷䛿 TaqMan Gene Expression Assay 䛾䜏䚸䛣䛾⏬㠃ୖ䛛䜙䛣䛾䜎䜎Ⓨὀసᴗ䛻㐍䜐䛣䛸䛜䛷䛝
䜎䛩䚹Ⓨὀ䛩䜛᫬䛻䛿 TaqMan Gene Expression Assay 䛻䝏䜵䝑䜽䜢ධ䜜䚸Add Selected to
Shopping List 䜢䜽䝸䝑䜽䛧䜎䛩䚹ὀグ䠖䛣䛾Ⓨὀ䛿䜲䞁䝍䞊䝛䝑䝖䜸䞊䝎䞊䝅䝇䝔䝮䜢฼⏝䛧䜎䛩䚹䜲䞁䝍䞊䝛䝑䝖䜸䞊䝎䞊䝅䝇䝔䝮䜢䛤฼⏝䛔䛯䛰䛟䛯䜑䛻䛿஦๓Ⓩ㘓䛜ᚲせ䛷䛩䚹

11d Experiment Shopping List ෆ䛻Ⓨὀ䛩䜛 TaqMan Gene Expression Assay 䛜䛒䜛䛣䛸䜢☜ㄆ䛧䛯䜙䚸
Order Materials in List 䜢䜽䝸䝑䜽䛧䜎䛩䚹

11a
11b

11c

11d

11e Order Materials – Log In ⏬㠃䛜⾲♧䛥䜜䜛䛾䛷䚸䜲䞁䝍䞊䝛䝑䝖䜸䞊䝎䞊䝅䝇䝔䝮⏝䛾 User Name 䛸Password 䜢ධຊ䛧䜎䛩䚹䛣䛾ඛ䛿㏻ᖖ䛾䜲䞁䝍䞊䝛䝑䝖䜸䞊䝎䞊䛸ྠᵝ䛻సᴗ䜢㐍䜑䜎䛩䚹඲䛶䛾సᴗ䛜⤊஢䛧䛯䜙 Finish Designing Experiment 䜢䜽䝸䝑䜽䛧䜎䛩䚹- 14 -

JP-886̲本文̲0707se.indd 13

15/07/08 17:33

12. ᭱⤊ⓗ䛺タᐃ☜ㄆ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
12a 䝥䝺䞊䝖䛾タᐃ䜢䛤☜ㄆୗ䛥䛔䚹ὀグ䠖ಟṇ䛧䛯䛔䛸䛝䛻䛿 Return to the Wizard 䜢䜽䝸䝑䜽䛧䚸Design Wizard ⏬㠃䛻ᡠ䛳䛶タᐃኚ᭦䛧䜎䛩䚹12b タᐃ䛻ၥ㢟䛺䛡䜜䜀 Save Experiment 䜢䜽䝸䝑䜽䛧䜎䛩䚹

12a ὀ

12b

㻌㻌㻌㻌㻌㻌㻌
12c Save Experiment 䝎䜲䜰䝻䜾䝪䝑䜽䝇䛷 Save 䜢䜽䝸䝑䜽䛩䜛䛸䚸1A 䛷タᐃ䛧䛯 Experiment Name 䛜䝣䜯䜲䝹ྡ䛸䛧䛶䚸ึᮇタᐃ䛾䝣䜷䝹䝎ෆ䛻ಖᏑ䛥䜜䜎䛩䚹ὀグ䠖ึᮇタᐃ䛷䛿䚸䝣䜯䜲䝹䛿 Applied Biosystems¥StepOne System v2.X¥experiments 䝣䜷䝹䝎䛻ಖᏑ䛥䜜䜎䛩䚹䛺䛚ಖᏑඛ䛾䝣䜷䝹䝎䛿௵ព䛻タᐃ䞉ᣦᐃ䛷䛝䜎䛩䚹ึᮇタᐃ䛾ಖᏑඛ䜢ኚ᭦䛩䜛䛸䛝䛿 Tools > Preferences䜢㑅ᢥᚋ䚸General 䝍䝤䜢㑅ᢥ䛧䚸Default Data Folder 䛻௵ព䛾ሙᡤ䜢ᣦᐃ䛧䛶ୗ䛥䛔䚹12c㻌㻌㻌㻌㻌㻌㻌㻌
12d ಖᏑ䛜᏶஢䛩䜛䛸䝩䞊䝮⏬㠃䛻ᡠ䜚䜎䛩䚹- 15 -

JP-886̲本文̲0707se.indd 14

15/07/08 17:33

⿵㊊䠖సᴗ㏵୰䛷 Finish Designing Experiment 䜢䜽䝸䝑䜽䛧䛯䜚䚸Save 䜢䜻䝱䞁䝉䝹䛧䛯ሙྜ

Design Wizard సᴗ㏵୰䛷 Designing Experiment 䜢䜽䝸䝑䜽䛧䛯䜚䚸Save 䜢䜻䝱䞁䝉䝹䛩䜛䛸䚸Wizard 䛜⤊஢䛧䝣䜯䜲䝹䛜㛤䛔䛯≧ែ䛸䛺䜚䜎䛩䚹Design Wizard ⏬㠃䛿䚸ᮏ᮶䛣䛾䝣䜯䜲䝹ୖ䛷⾜䛖タᐃ䛾సᴗ䜢⡆␎໬䛧䛯䜒䛾䛷䛩䚹Design Wizard 䛷タᐃ䛷䛝䜛඲䛶䛾㡯┠䛿䚸䛣䛾䝣䜯䜲䝹ୖ䛷┤᥋タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹⏬㠃ᕥഃ䛾 Setup ෆ䛾ྛ㡯┠䜢㑅ᢥ䛩䜛䛸䚸ྑഃ䛻㑅ᢥ䛧䛯㡯┠䛾タᐃ⏬㠃䛜⾲♧䛥䜜䜎䛩䛾䛷䚸௵ព䛻タᐃ䞉ኚ᭦䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹タᐃ᪉ἲ䛾ヲ⣽䛻䛴䛔䛶䛿䚸ḟ䝨䞊䝆௨㝆䜢䛤ཧ↷䛟䛰䛥䛔䚹- 16 -

JP-886̲本文̲0707se.indd 15

15/07/08 17:33

Standard Curve Method ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ
StepOne 䝋䝣䝖䜴䜵䜰䛾 Advanced Wizard 䜢⏝䛔䛶᪂つᐇ㦂⣔䜢సᡂ䛧䜎䛩䚹

1.

StepOne 䝋䝣䝖䜴䜵䜰䛾䝅䝵䞊䝖䜹䝑䝖䜢䝎䝤䝹䜽䝸䝑䜽䛧䛶䚸䝋䝣䝖䜴䜵䜰䜢㉳ື䛧䜎䛩䚹ὀグ䠖StepOne 䝋䝣䝖䜴䜵䜰䛾㉳ື䛻䛿⣙ 15 ⛊⛬ᗘ䛛䛛䜚䜎䛩䚹

2.

Login ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹୍᫬ⓗ䛻䝻䜾䜲䞁䛩䜛䛸䛝䛻䛿 Log in as Guest 䜢䜽䝸䝑䜽䛧䜎䛩䚹䝴䞊䝄䞊Ⓩ㘓䛧䛶౑⏝䛩䜛ሙྜ䛿䚸᪂䛧䛔 User Name 䜢䝍䜲䝥ධຊ䛧䚸OK 䜢䜽䝸䝑䜽䛧䜎䛩䚹㻌ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹ὀグ䠖୍ᗘⓏ㘓䛧䛯䝴䞊䝄䞊ྡ䛿䚸ḟᅇ䛛䜙䝥䝹䝎䜴䞁䝯䝙䝳䞊䜘䜚㑅ᢥ䛷䛝䜛䜘䛖䛻䛺䜚䜎䛩䚹3.

䝩䞊䝮⏬㠃䛜⾲♧䛥䜜䜎䛩䚹
Design Wizard 䛾ୗ䛾▮༳䝪䝍䞁䜢䜽䝸䝑䜽䛧䜎䛩䚹- 17 -

JP-886̲本文̲0707se.indd 16

15/07/08 17:33

4.

䝪䝍䞁䛜㏣ຍ⾲♧䛥䜜䜛䛾䛷䚸䛭䛾୰䛾 Advanced Setup 䜢䜽䝸䝑䜽䛧䜎䛩䚹5.

᪂つᐇ㦂⣔䝣䜯䜲䝹䛜㛤䛝䜎䛩䚹

B

AC
A : Experiment Menuᐇ㦂䜢⾜䛖䛸䛝䛻䛿䚸䛣䛾䝯䝙䝳䞊䜢ୖ䛛䜙㡰␒䛻ษ䜚᭰䛘䜛䛣䛸䛷䝗䜻䝳䝯䞁䝖䛾タᐃ䚸䝷䞁䛚䜘䜃ゎᯒ䜎䛷⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹B : Experiment Menu 䛷㑅ᢥ䛧䛯㡯┠䛻ᑐᛂ䛩䜛⏬㠃䛜⾲♧䛥䜜䜎䛩䚹C : 䝣䜯䜲䝹䜢㛤䛟䛸⮬ືⓗ䛻䝋䝣䝖䜴䜵䜰ୗẁ䛻䝍䝤䛜⾲♧䛥䜜䜎䛩䚹䝍䝤䜢䜽䝸䝑䜽䛩䜛䛣䛸䛷䚸⾲♧䛩䜛䝣䜯䜲䝹䜢ษ䜚᭰䛘䛯䜚䚸Home ⏬䛻ᡠ䜛䛣䛸䛜䛷䛝䜎䛩䚹ྛ䝣䜯䜲䝹䜢㛢䛨䜛䛸䛝䛻䛿䚸䝍䝤ྑഃ䛾 X 䜢䜽䝸䝑䜽䛧䜎䛩䚹

- 18 -

JP-886̲本文̲0707se.indd 17

15/07/08 17:33

6.

Experiment Menu 䛾 Experiment Properties 䜢䜽䝸䝑䜽䛧䜎䛩䚹
6a

How do you want to identify this experiment?
Experiment Name 䛾ධຊ䛿ᚲ㡲䛷䛩䚹䛣䛾ྡ๓䛜䛭䛾䜎䜎䝣䜯䜲䝹ྡ䛻䛺䜚䜎䛩䚹䛭䛾䜋䛛䛾㡯┠䛾ධຊ䛿௵ព䛷䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹

6bWhich instrument are you using to run the experiment?
StepOnePlus (96well) 䜎䛯䛿 StepOne (48well) ఱ䜜䛛䛤౑⏝䛾ᶵჾ䜢㑅ᢥ䛧䛶䛟䛰䛥䛔䚹ὀグ䠖㛫㐪䛳䛯ᶵჾ䜢㑅ᢥ䛧䛯ሙྜ཯ᛂ䜢ጞ䜑䜛䛣䛸䛜䛷䛝䜎䛫䜣䚹6cWhat type of experiment do you want to set up?
ᐃ㔞䜰䝑䝉䜲䜢⾜䛖䛸䛝䛻䛿 Quantitation 䛾䛥䜙䛻䛹䛾ᡭἲ䛷⾜䛖䛛㑅ᢥ䛧䜎䛩䚹
Standard Curve : ཯ᛂ⣔ẖ䛻᳨㔞⥺䜢సᡂ䛧䚸ᐃ㔞್䜢⟬ฟ䛧䜎䛩
Relative Standard Curve : ཯ᛂ⣔ẖ䛻᳨㔞⥺䛛䜙ᐃ㔞್䜢ồ䜑䛯䛒䛸䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛾್䛷䛾⿵ṇཬ䜃䜻䝱䝸䝤䝺䞊䝍䝃䞁䝥䝹䛸䛾┦ᑐ್䜢䚸䝋䝣䝖䜴䜵䜰䛜⮬ືⓗ䛻⟬ฟ䛧䜎䛩䚹Comparative CT (ǻǻCT) : ẚ㍑ CT ἲ䛷䛾┦ᑐᐃ㔞䜢䝋䝣䝖䜴䜵䜰䛜⮬ືⓗ䛻⟬ฟ䛧䜎䛩䚹6dWhich reagents do you want to use to detect the target sequence?
཯ᛂ⣔䛾䜿䝭䝇䝖䝸䜢㑅ᢥ䛧䜎䛩䚹㻌 SYBR Green Reagents 䜢㑅ᢥ䛩䜛䛸ᕥୗ䛻 Include Melt Curve䝏䜵䝑䜽䝪䝑䜽䝇䛜⾲♧䛥䜜䜎䛩䚹6eWhich ramp speed do you want to use in the instrument run?
䝷䞁䝰䞊䝗䜢㑅ᢥ䛧䜎䛩䚹䛤฼⏝䛔䛯䛰䛟ヨ⸆䛻ྜ䜟䛫䛶 Standard 䜎䛯䛿 Fast 䛛䜙㑅ᢥ䛧䜎䛩䚹

6a

6b

6c
6d
6e

- 19 -

JP-886̲本文̲0707se.indd 18

15/07/08 17:33

7.

Experiment Menu 䛾 Pate Setup 䜢䜽䝸䝑䜽䛧䜎䛩䚹䛣䛾⏬㠃䛻䛿䇿Define Targets and Samples䇿䛸䇿Assign Targets and Samples䇿䛾 2 䛴䛾䝍䝤䛜䛒䜚䜎䛩䚹ึᮇタᐃ䛷䛿䛿䛨䜑䛻 Define Targets and Samples 䝍䝤⏬㠃䛜⾲♧䛥䜜䜎䛩䚹

Define Targets and Samples 䝍䝤⏬㠃ᕥഃ䛾 Define Targets 䛷 Target ᝟ሗ䜢ධຊ䛧䜎䛩7a䝍䞊䝀䝑䝖䠄䝥䝷䜲䝬䞊䝉䝑䝖䠅䛾ᩘ䛰䛡 Add New Target 䜢䜽䝸䝑䜽䛧䛷 Target 䜢㏣ຍ䛧䜎䛩䚹䐟Target Name 䛻㑇ఏᏊྡ䜢ධຊ䛧䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹䐠

Reporter 䛷䛿᳨ฟ䛩䜛⺯ගⰍ⣲䜢㑅ᢥ䛧䜎䛩䚹䐡Quencher 䛷䛿䚸TaqMan 䝥䝻䞊䝤䛾ሙྜ䛿”TAMRA”䚸TaqMan MGB 䝥䝻䞊䝤䛾ሙྜ䛿”NFQ-MGB”䚸SYBR Green 䛾ሙྜ䛿”None”䜢㑅ᢥ䛧䜎䛩䚹ὀグ䠖StepOne 䝅䝇䝔䝮䛷䛿䚸TAMRA Ⰽ⣲䜢⏝䛔䛯ᐇ㦂䜢⾜䛖䛣䛸䛿䛷䛝䜎䛫䜣䚹7bసᡂ䛧䛯 Target 䛿 Library 䛻ಖᏑ䛧䛶⧞䜚㏉䛧౑⏝䛩䜛䛣䛸䛜ฟ᮶䜎䛩䚹䐟ಖᏑ䛧䛯䛔 Target 䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖䛥䜜䛯䜙䚸Save Target 䝪䝍䞁䜢䜽䝸䝑䜽䛩䜛䛸☜ㄆ⏬㠃䛜⾲♧䛥䜜䛶䛛䜙ಖᏑ䛥䜜䜎䛩䚹䐠

Library 䛻ಖᏑ䛧䛯 Target 䜢࿧䜃ฟ䛧䛶౑⏝䛧䛯䛔ሙྜ䛿䚸Add Saved Target 䜢䜽䝸䝑䜽䛧 Target
Library 䜢⾲♧䛧䜎䛩䚹䐡౑⏝䛩䜛 Target 䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖䛥䜜䛯䜙 Add Selected Target(s)䜢䜽䝸䝑䜽䛩䜛䛸䝸䝇䝖䛻㏣ຍ䛥䜜䜎䛩䚹
7a

7b䐟

7b䐠
7

7a䐟

7a䐠

7a䐡

7b䐡

- 20 -

JP-886̲本文̲0707se.indd 19

15/07/08 17:33

Define Targets and Sample 䝍䝤ෆ䛾 Define Samples 䛷 Sample ᝟ሗ䜢ධຊ䛧䜎䛩䚹7cᮍ▱䝃䞁䝥䝹䛾ᩘ䛰䛡 Add New Samples 䜢䜽䝸䝑䜽䛧䛶 Sample 䜢㏣ຍ䛧䜎䛩䚹

7d

Sample Name 䛻ྛ䝃䞁䝥䝹䛾ྡ๓䜢ධຊ䛧䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹7eసᡂ䛧䛯 Sample 䛿 Library 䛻ಖᏑ䛧䛶⧞䜚㏉䛧౑⏝䛩䜛䛣䛸䛜ฟ᮶䜎䛩䚹䐟ಖᏑ䛧䛯䛔 Sample 䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖䛥䜜䛯䜙䚸Save Sample 䝪䝍䞁䜢䜽䝸䝑䜽䛩䜛䛸☜ㄆ⏬
㠃䛜⾲♧䛥䜜䛶䛛䜙ಖᏑ䛥䜜䜎䛩䚹䐠

Library 䛻ಖᏑ䛧䛯 Sample 䜢࿧䜃ฟ䛧䛶౑⏝䛧䛯䛔ሙྜ䛿䚸Add Saved Sample 䜢䜽䝸䝑䜽䛧Sample Library 䜢⾲♧䛧䜎䛩䚹䐡౑⏝䛩䜛 Sample 䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖䛥䜜䛯䜙 Add Selected Sample(s)䜢䜽䝸䝑䜽䛩䜛䛸䝸䝇䝖
䛻㏣ຍ䛥䜜䜎䛩䚹
7c

7e䐟

7e䐠
7d

7e䐡

- 21 -

JP-886̲本文̲0707se.indd 20

15/07/08 17:33

Assign Targets and Samples 䝍䝤䜢䜽䝸䝑䜽䛧⾲♧䛧䜎䛩䚹ྛ䜴䜵䝹䛻 Target 䛸 Task䚸Sample 䜢タᐃ䛧䜎䛩䚹
7f

View Plate Layout ෆ䛷䚸タᐃ䛩䜛䜴䜵䝹䜢㑅ᢥ䛧䜎䛩䚹- 䝬䜴䝇䜢䝗䝷䝑䜾䛥䛫䜛䛸㑅ᢥᯟ䛜ᗈ䛜䜚䜎䛩䚹- Control 䜻䞊䜢ᢲ䛧䛺䛜䜙䜽䝸䝑䜽䛩䜛䛸䚸௵ព䛾䜴䜵䝹䜢㑅ᢥ䛧䛯䜚๐㝖䛧䛯䜚䛷䛝䜎䛩䚹

7g

Assign target(s) to the selected wells 䝣䜱䞊䝹䝗ෆ䛾䝍䞊䝀䝑䝖䝸䝇䝖䛾 Assign 䛻䝏䜵䝑䜽䜢ධ䜜䜛䛸䚸㑅ᢥ䛧䛯䜴䜵䝹䛻 Target 䛜タᐃ䛥䜜䜎䛩䚹䐟

Task 䜢㑅ᢥ䛧䜎䛩䚹ึᮇタᐃ䛷䛿඲䛶”U”䛷䛩䚹䜰䝹䝣䜯䝧䝑䝖䜢䜽䝸䝑䜽䛧䛶タᐃ䜢㐺ᐅኚ᭦䛧䜎䛩䚹᳨㔞⥺䜢సᡂ䛧䛶ᐃ㔞䜢⾜䛖㝿䛻䛿䚸ୗ䛾 3 ✀䛾䝍䝇䜽䜢タᐃ䛧䜎䛩䚹㻌 U : Unknown䠙ᐃ㔞䜢⾜䛔䛯䛔ᮍ▱䝃䞁䝥䝹㻌 S : Standard䠙᳨㔞⥺సᡂ⏝ᶆ‽䝃䞁䝥䝹㻌 ὀ䠖Standard 䛾タᐃ䛿 Define and Set Up Standards 䛾ᶵ⬟䠄ḟ䝨䞊䝆ཧ䠅䜢౑⏝䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹㻌 N : No Template Control䠙㗪ᆺ䜢ྵ䜎䛺䛔཯ᛂ䜴䜵䝹䐠

Quantity 䛻 Standard䠄᳨㔞⥺సᡂ⏝ᶆ‽䝃䞁䝥䝹䠅䛾ึᮇ㔞䜢ධຊ䛧䜎䛩䚹ὀ䠖 Quantity 䛾ධຊ䛿 Define and Set Up Standards 䛾ᶵ⬟䠄ḟ䝨䞊䝆ཧ䠅䜢౑⏝䛩䜛䛣䛸䛷⮬ື໬䛷䛝䜎䛩䚹7hUnknown 䛻 Sample 䜢タᐃ䛧䜎䛩䚹タᐃ䛩䜛 Unknown 䜴䜵䝹䜢㑅ᢥ䛧䚸Assign sample(s) to theselected wells 䝣䜱䞊䝹䝗ෆ䛾ヱᙜ䝃䞁䝥䝹䛾 Assign 䛻䝏䜵䝑䜽䜢ධ䜜䜛䛸㑅ᢥ䛧䛯䜴䜵䝹䛻 Sample 䛜タᐃ䛥䜜䜎䛩䚹

7i

Show in Wells 䝪䝍䞁䜢䜽䝸䝑䜽䛧䛶䜴䜵䝹䛾⾲♧タᐃ䜢ኚ᭦䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

7j

StepOnePlus 䛾ሙྜ䚸VeriFlexTM ᶵ⬟䜢⏝䛔䛶௵ព䛾䝇䝔䞊䝆䛷ྛ䝤䝻䝑䜽

ᗘ䜢␗䛺䜛

ᗘ䛷タᐃ䛩

䜛䛣䛸䛜ฟ᮶䜎䛩䚹(59 䝨䞊䝆ཧ)

7j
7i
7g

7g䐟

7f

7g䐠

7g䐠ὀ
7h

㻌
- 22 -

JP-886̲本文̲0707se.indd 21

15/07/08 17:33

Define and Set up Standards 䜢⏝䛔䛯 Standard 䛾タᐃ᪉ἲ⏬㠃ᕥ୰ኸ䛾 Define and Set up Standards 䝪䝍䞁䜢䜽䝸䝑䜽䛧䛶⾲♧䛥䜜䜛⏬㠃䛷䚸Standard 䛾タᐃ䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹
iStandard 䜢タᐃ䛩䜛 Target 䜢䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛧䜎䛩䚹iiStandard 䛻タᐃ䛩䜛⃰ᗘ䛾ᩘ䜢ධຊ䛧䜎䛩䚹㻌 㻌 㻌 -㻌 ⃰ᗘ䛾ᩘ䛿 5 Ⅼ௨ୖタᐃ䛩䜛䛣䛸䜢᥎ዡ䛧䜎䛩䚹᭱ప䛷䜒 3 Ⅼ䛿䛤⏝ពୗ䛥䛔䚹iii

ྛ Standard 䛾཯᚟ᩘ䜢ධຊ䛧䜎䛩䚹iv

Standard 䛾⃰ᗘ䛾䚸ᇶⅬ䛸䛺䜛್䜢ධຊ䛧䜎䛩䚹vᕼ㔘ಸ⋡䜢䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙㑅ᢥ䛧䜎䛩䚹㻌 㻌 㻌 - 1:5 䜢㑅ᢥ䛩䜛䛸䚸ᇶⅬ䛾⃰ᗘ䛻ᑐ䛧䛶 1/5 䛾್䛜⧞䜚㏉䛧⮬ືⓗ䛻⟬ฟ䛥䜜䜎䛩䚹㻌 㻌 㻌 - X10 䜢㑅ᢥ䛩䜛䛸䚸ᇶⅬ䛾⃰ᗘ䛻ᑐ䛧䛶 10 ಸ䛾್䛜⧞䜚㏉䛧⮬ືⓗ䛻⟬ฟ䛥䜜䜎䛩䚹vi䜴䜵䝹䛾⮬ື㓄⨨䛾タᐃ䜢⾜䛔䜎䛩䚹㻌 㻌 㻌 - Automatically Select Wells for Me 䜢㑅ᢥ䛩䜛䛸⮬ືⓗ䛻䜴䜵䝹䜢Ỵᐃ䛧䜎䛩䚹㻌 㻌 㻌 - Let Me Select Wells 䜢㑅ᢥ䛩䜛䛸䚸௵ព䛾఩⨨䜢⮬ศ䛷㑅ᢥ䛧䛶タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹vii

Standard 䛜タᐃ䛥䜜䜛䜴䜵䝹䛾⠊ᅖ䛜䝝䜲䝷䜲䝖䛷⾲♧䛥䜜䜎䛩䚹vi 䛷 Let Me Select Wells 䜢㑅ᢥ䛧䛯᫬䛻䛿䚸䛣䛾ሙᡤ䛷タᐃ䛩䜛䜴䜵䝹䜢㑅ᢥ䛧䜎䛩䚹viiiᕼ㔘⣔ิ䛾୪䜃㡰䛾タᐃ䛷䛩䚹Rows 䜢㑅ᢥ䛩䜛䛸ᶓ᪉ྥ䛻䚸Columns 䜢㑅ᢥ䛩䜛䛸⦪᪉ྥ䛻䚸䛭䜜䛮䜜୪䜃䜎䛩䚹

ix

Apply 䜢䜽䝸䝑䜽䛩䜛䛸䚸ୖグタᐃ䛻ᚑ䛳䛶䚸䜴䜵䝹䜈䛾 Standard Sample 䛾タᐃ䛜ᐇ⾜䛥䜜䜎䛩䚹x」ᩘ䛾 Target 䛜䛒䜛䛸䛝䛻䛿䚸I 䛛䜙 iX 䜎䛷䛾సᴗ䜢⧞䜚㏉䛧䜎䛩䚹タᐃ䛜⤊஢䛧䛯䜙 Close 䜢䜽䝸䝑䜽䛧䚸⏬㠃䜢㛢䛨䜎䛩䚹

i
ii
iii
iv
v

vi

vii
x

viii
ix

- 23 -

JP-886̲本文̲0707se.indd 22

15/07/08 17:33

8.

Graphical View 䝍䝤䛛 Tabular View 䝍䝤䜢㑅ᢥ䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Graphical View 䝍䝤⏬㠃䜢⏝䛔䛶ㄝ᫂䛧䜎䛩䛜䚸䛹䛱䜙䛾⏬㠃䛷䜒ྠ䛨సᴗ䛜䛷䛝䜎䛩䛾䛷䚸ぢ䜔䛩䛔⏬㠃䜢㑅ᢥ䛧䛶ୗ䛥䛔䚹

8a཯ᛂ䝪䝸䝳䞊䝮䜢ධຊ䛧䜎䛩䚹StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛷䛿 10-30μL 䛾⠊ᅖෆ䛷ᐇ㦂䜢⾜䛖䛣䛸䜢᥎ዡ䛧䛶䛔䜎䛩䚹ὀグ䠖ᐇ㦂ྍ⬟䛺᭱኱䝪䝸䝳䞊䝮䛿 50μL 䛷䛩䚹31μL ௨ୖ䛻タᐃ䛩䜛䛸䝷䞁䝥䝇䝢䞊䝗䛜㐜䛟䛺䜚䜎䛩8b䝃䞊䝬䝹䝃䜲䜽䝷䞊᮲௳䜢☜ㄆ䛧䜎䛩䚹ᚲせ䛻ᛂ䛨䛶䚸ᕥୖ䛾䝪䝍䞁䜢㑅ᢥ䛩䜛䛣䛸䛷䚸௵ព䛾䝥䝻䜾䝷
䝮䜢㏣ຍ䞉๐㝖䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹8c䛾䝬䞊䜽䛜䜰䜽䝔䜱䝤䛻䛺䛳䛶䛔䜛䝫䜲䞁䝖䛷 StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛿⺯ග䝕䞊䝍䜢ᅇ཰䛧䜎䛩䚹䝬䞊䜽䛾ୖ䜢䜽䝸䝑䜽䛩䜛䛣䛸䛷䚸䝕䞊䝍ᅇ཰䝫䜲䞁䝖䛾ᩘ䜢タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹ὀグ䠖䝃䜲䜽䝹䝇䝔䞊䝆ෆ䛷䛾䝕䞊䝍ᅇ཰䝫䜲䞁䝖䛿ᚲ䛪 1 䞄ᡤ䛻䛺䜛䜘䛖䛻䛤ὀព䛟䛰䛥䛔䚹ὀグ䠖䝕䞊䝍ᅇ཰䝫䜲䞁䝖䛾䝬䞊䜽䛜䚸䝃䜲䜽䝹䝇䝔䞊䝆䛾᭱ᚋ䛾䝇䝔䝑䝥䛷䜰䜽䝔䜱䝤䛻䛺䛳䛶䛔䜛䛣䛸䜢䛤☜ㄆ䛟䛰䛥䛔䚹䛣䛾䝬䞊䜽䛿 1 ᅇ䜽䝸䝑䜽䛩䜛䛸㠀䜰䜽䝔䜱䝤䛻䛺䜚䚸䛭䛾䜎䜎䝷䞁䜢⾜䛖䛸䝕䞊䝍ᅇ཰䛜⾜䜟䜜䜎䛫䜣䚹

8d

Run Method 䛾᮲௳䜢ಖᏑ䛩䜛ሙྜ䛻䛿 Save Run Method 䜢䜽䝸䝑䜽䛧䜎䛩䚹Name 䜢㐺ᐅධຊ䛧䚸
Save 䜢䜽䝸䝑䜽䛩䜛䛸ಖᏑ䛥䜜䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹8eಖᏑ䛧䛯 Run Method 䛿䚸ḟᅇ௨㝆ྠ䛨᮲௳䛷ᐇ㦂䜢⾜䛖䛸䛝䛻 Open Run Method 䜢䜽䝸䝑䜽䛧䜎䛩䚹⾲♧䛥䜜䛯 Select Run Method ⏬㠃ୖ䛷௵ព䛾 Run Method 䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖㑅ᢥ䛧䚸OK 䝪䝍䞁䜢䜽䝸䝑䜽䛧䜎䛩䚹

8a
8d
8b
8e

8c

- 24 -

JP-886̲本文̲0707se.indd 23

15/07/08 17:33

9.

䝣䜯䜲䝹䛾ಖᏑ
䝷䞁䜢㛤ጞ䛩䜛䜎䛘䛻䚸ᚲ䛪䝣䜯䜲䝹䜢ಖᏑ䛩䜛䛣䛸䛜ᚲせ䛷䛩䚹

9a⏬㠃ᕥୖ䛾 Save 䜢䜽䝸䝑䜽䛧䜎䛩䚹9bಖᏑඛ䛾ᣦᐃ䛾⏬㠃䛜⾲♧䛥䜜䜎䛩䚹ಖᏑඛ䜢ᣦᐃ䛧䚸File Name 䜢☜ㄆ䛧䚸Save 䜢䜽䝸䝑䜽䛩䜛䛸ಖᏑ䛥䜜䜎䛩䚹ὀグ䠖ึᮇタᐃ䛷䛿䚸䝣䜯䜲䝹䛿 Applied Biosystems¥ StepOne Software v2.X¥experiments 䝣䜷䝹䝎䛻ಖᏑ䛥䜜䜎䛩䚹䛺䛚ಖᏑඛ䛾䝣䜷䝹䝎䛿௵ព䛻タᐃ䞉ᣦᐃ䛷䛝䜎䛩䚹C:䝗䝷䜲䝤䜈䛾ಖᏑ䛿㑊䛡䚸D:䝗䝷䜲䝤䛻ಖᏑ䜢⾜䛳䛶ୗ䛥䛔䚹ึᮇタᐃ䛾ಖᏑඛ䜢ኚ᭦䛩䜛䛸䛝䛿 Tools > Preferences 䜢㑅ᢥᚋ䚸Defaults 䝍䝤䜢㑅ᢥ䛧䚸Default Data
Folder 䛻௵ព䛾ሙᡤ䜢ᣦᐃ䛧䛶ୗ䛥䛔䚹9c䠄䜸䝥䝅䝵䞁䠅⧞䜚㏉䛧ྠ䛨᮲௳䛷ᐇ㦂䜢⾜䛖ሙྜ䛻䚸Template 䝣䜯䜲䝹䜢సᡂ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹䛣䛾ሙྜ䛿⏬㠃ᕥୖ䛾 Save 䛾ᶓ䛾䕰䜢䜽䝸䝑䜽䛧䚸Save as Template 䜢㑅ᢥ䛧䜎䛩䚹ಖᏑඛ䜢ᣦᐃ䛧䚸File Name 䜢ධ䜜䛶䚸Save 䜢䜽䝸䝑䜽䛩䜛䛸ಖᏑ䛥䜜䜎䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹సᡂ䛧䛯 Template 䝣䜯䜲䝹䛛䜙᪂つ䝣䜯䜲䝹䜢సᡂ䛩䜛䛸䛝䛻䛿䚸䝋䝣䝖䜴䜵䜰㉳ືᚋึ䜑䛻⾲♧䛥䜜䜛
Home ⏬㠃䛷䚸Setup ෆ Template 䝪䝍䞁䜢䜽䝸䝑䜽䛧䚸Template 䝣䜯䜲䝹䜢㑅ᢥ䛧䛶㛤䛝䜎䛩䚹ὀグ䠖Template 䝣䜯䜲䝹䛿 D:¥Applied Biosystems¥StepOne Software v2.X¥config¥templates 䝣䜷䝹䝎ෆ䛻ಖᏑ䛩䜛䛸䚸Home ⏬㠃䛷 Setup ෆ Template 䝪䝍䞁䜢䜽䝸䝑䜽䛧䛯᫬䛻䚸䛩䛠䛻 Template 䝣䜯䜲䝹䛜⾲♧䛥䜜䜎䛩䚹ὀグ䠖Template 䝣䜯䜲䝹䛿ᚲ䛪䝷䞁๓䛾䝣䜯䜲䝹䜢ಖᏑ䛧䛶సᡂ䛧䛶ୗ䛥䛔䚹䝷䞁䛜⤊䜟䛳䛯䛒䛸䛾䝣䜯䜲䝹䛛䜙ูྡಖᏑ䛷 Template 䝣䜯䜲䝹䜢ಖᏑ䛩䜛䛸䚸䝕䞊䝍ᅇ཰䛜ṇᖖ䛻⾜䜟䜜䛺䛔➼䝖䝷䝤䝹䛜Ⓨ⏕䛩䜛༴㝤ᛶ䛜䛒䜚䜎䛩䚹- 25 -

JP-886̲本文̲0707se.indd 24

15/07/08 17:33

཯ᛂ䝥䝺䞊䝖䛾‽ഛ᥎ዡ䛾ᾘ⪖ရཬ䜃⤌䜏ྜ䜟䛫䛿ୗグ䛾㏻䜚䛷䛩䚹㔜せ䠖཯ᛂ⣔ㄪᩚ⏝䛾ᾘ⪖ရ䜢ྲྀ䜚ᢅ䛖㝿䛻䛿ᚲ䛪䝟䜴䝎䞊䝣䝸䞊䛾ᡭ⿄䜢⿦╔䛧䛶ୗ䛥䛔䚹㔜せ䠖཯ᛂ䛩䜛䝥䝺䞊䝖䜔䝏䝳䞊䝤➼䛻䝬䝆䝑䜽䛷䝷䝧䝹䜢䛧䛺䛔䛷䛟䛰䛥䛔䚹䝬䝆䝑䜽䛾୰䛻䛿⺯ග≀㉁䛜ྵ䜎䜜䛶䛔䜛䛯䜑䚸䝕䞊䝍䛻ᝏᙳ㡪䜢ཬ䜌䛧䜎䛩䚹

StepOne System
ၟရྡMicroAmp Fast Optical 48-Well Reaction Plates
MicroAmp Optical 48-Well Adhesive Film
MicroAmp Fast 8-Tube Strips
MicroAmp Optical 8-Cap Strips
MicroAmp Fast Reaction Tubes with Caps
MicroAmp Fast 48-Well Trays
MicroAmp 48-Well Base Adaptor
MicroAmp 96-Well Support Base
MicroAmp Adhesive Film Applicatorၟရ␒ྕ
43758164375323435829343230324358297437528243752844379590
4333183

ᅗ
A
G
E
D
F
B
H
C
-

ὀグ䠖8 㐃䜻䝱䝑䝥䛿ᚲ䛪 Flat 䝍䜲䝥䜢䛤฼⏝ୗ䛥䛔䚹㔜せ䟿䠖䝥䝺䞊䝖௨እ䜢฼⏝䛩䜛㝿䛿䚸4 ᮏ௨ୖ䛾䝏䝳䞊䝤䜢䝖䝺䜲䛻䝃䞁䝥䝹䝤䝻䝑䜽୰ᚰ䜘䜚௜㏆䜘䜚䝉䝑䝖䛧䛶䛟䛰䛥䛔䚹

StepOnePlus System
ၟရྡMicroAmp Fast 96-Well Reaction Plate
MicroAmp Optical Adhesive Film
MicroAmp Fast 8-Tube Strips
MicroAmp Optical 8-Cap Strips
MicroAmp Fast Reaction Tubes with Caps
MicroAmp 96-Well Tray for VeriFlexTM Blocks
MicroAmp 96-Well Support Base
MicroAmp Adhesive Film Applicatorၟရ␒ྕ
4346907
4311971435829343230324358297
43799834379590
4333183

ᅗ
A
G
E
D
F
B
C
-

ὀグ䠖8 㐃䜻䝱䝑䝥䛿ᚲ䛪 Flat 䝍䜲䝥䜢䛤฼⏝ୗ䛥䛔䚹㔜せ䟿䠖䝥䝺䞊䝖௨እ䜢฼⏝䛩䜛㝿䛿䚸16 ᮏ௨ୖ䛾䝏䝳䞊䝤䜢䝖䝺䜲䛻⦪䠄A-H䠅2 ิ䜎䛯䛿ᶓ䠄3-10䠅2 ิ䛻୪䜉䛶䝃䞁䝥䝹䝤䝻䝑䜽୰ᚰ䜘䜚䝉䝑䝖䛧䛶䛟䛰䛥䛔䚹- 26 -

JP-886̲本文̲0707se.indd 25

15/07/08 17:33

Optical Reaction Plate 䛻 Adhesive Film 䜢㈞䜛᪉ἲ

1.

Optical Reaction Plate 䜢 96-Well㻌 base 䛾୰ኸ䛻䝉䝑䝖䛧䜎䛩䚹48-Well Optical Reaction Plate 䛿
96-well Base 䛾ษ㎸䜏䛜ධ䛳䛶䛔䜛㒊ศ䛻ྜ䜟䛫䜛䛸䚸䛧䛳䛛䜚䛸䛿䜑㎸䜐䛣䛸䛜䛷䛝䜎䛩䚹㻌 㻌㻌 㻌 㻌 㻌 StepOne System

2.

StepOnePlus System㻌 㻌

Optical Adhesive Film 䜢୍ᯛྲྀ䜚ฟ䛧䚸䝭䝅䞁┠䛻ἢ䛳䛶ᢡ䜚᭤䛢䜎䛩䚹

3.

Film 䜢⿬㏉䛧䚸୰ኸ㒊ศ䜢ゐ䜙䛺䛔䜘䛖䛻䚸ⓑ䛔⿬⣬䜢ྲྀ䜚እ䛧䜎䛩䚹

4.

୧➃䛾䝍䝤䜢ᣢ䛱䚸Reaction Plate䛾ୖ䛻 Optical Adhesive Film 䜢䜖䛳䛟䜚䛸㍕䛫䜎䛩䚹

5.

Applicator 䜢⏝䛔䛶 Optical
Adhesive Film ୖ䜢୎ᑀ䛻䛺䛮䜚䚸
Cover 䛸 Plate 䛾⦕㒊ศ䜢ᐦ╔䛥䛫䜎䛩䚹

6.

Applicator 䛾➃䛷 Cover 䛾➃䜢ᢲ䛥䛘䚸䝍䝤䛾䜋䜌୰ኸ䜢ᣢ䛳䛶㍍䛟ᘬ䛔䛶ษ䜚㞳䛧䜎䛩䚹7.

䝇䝔䝑䝥 5 䜢䛥䜙䛻⾜䛔䜎䛩䚹㻌 ⦪᪉ྥཬ䜃ᶓ᪉ྥ䛻䚸䝥䝺䞊䝖ഃ䛻 Applicator 䜢ᢲ䛧௜䛡䛺䛜䜙⧞䜚㏉䛧䛣䛩䜚䜎䛩䚹

8.

᭱ᚋ䛻 Applicator 䛾䜶䝑䝆䛾㒊ศ䜢䛴䛛䛳䛶䚸䜴䜵䝹䛾እ࿘ᅖ䛻ἢ䛳䛶䛺䛮䜚䜎䛩䚹䛣䛾᫬䜒 Applicator䜢䝥䝺䞊䝖ഃ䛻䛧䛳䛛䜚䛸ᢲ䛧௜䛡䛶䛣䛩䜚䚸䝅䞊䝹䜢᏶඲䛻ᐦ╔䛥䛫䛶䛟䛰䛥䛔䚹

9.

᭱ᚋ䛻䝅䞊䝹䛜䜴䜵䝹ୖ㒊䛾⦕䛻ᐦ╔䛧䛶䛔䜛䛣䛸䜢☜ㄆ䛧䛶䛟䛰䛥䛔䚹- 27 -

JP-886̲本文̲0707se.indd 26

15/07/08 17:33

཯ᛂᾮ䜢䝏䝳䞊䝤ཪ䛿䝥䝺䞊䝖䛻ศὀ䛧䝅䞊䝹䜢䛧䛯䜙䚸䝥䝺䞊䝖㐲ᚰᶵ䛷㐲ᚰ䛧䚸ᚲ䛪཯ᛂᾮ䛜䜴䜵䝹䛾ᗏ䛻㞟䜎䛳䛶䛔䜛䛛䛤☜ㄆ䛟䛰䛥䛔䚹཯ᛂᾮ䛜䜴䜵䝹䛾ᗏ䛻㞟䜎䛳䛶䛔䛺䛔ሙྜ䛿㐲ᚰ䛧䛶䛟䛰䛥䛔䚹ὀグ䠖཯ᛂᾮ䛾Ỉ㠃䛻ᾋ䛛䜣䛷䛔䜛Ἳ䛿䛭䛾䜎䜎䛷䜒኱୔ኵ䛷䛩䚹䝷䞁㛤ጞᚋ䚸⇕ኚᛶ䛾䝇䝔䝑䝥䛷Ἳ䛿๭䜜䛶ᾘ䛘䜎䛩䚹- 28 -

JP-886̲本文̲0707se.indd 27

15/07/08 17:33

䝷䞁䛾㛤ጞ䛸⤊஢཯ᛂ䝥䝺䞊䝖䛾‽ഛ䛜䛷䛝䛯䜙䚸ಖᏑ䛧䛯ᐇ㦂᮲௳䝣䜯䜲䝹䜢㛤䛝䚸䝷䞁䜢㛤ጞ䛧䜎䛩䚹

1.

ಖᏑ䛧䛯ᐇ㦂᮲௳䝣䜯䜲䝹䜢㛤䛝䜎䛩䚹1a䜒䛧䝋䝣䝖䜴䜵䜰䛜⤊஢䛧䛶䛔䛯䛺䜙䚸StepOne 䝋䝣䝖䜴䜵䜰䛾䝅䝵䞊䝖䜹䝑䝖䜢䝎䝤䝹䜽䝸䝑䜽䛧䛶䚸䝋䝣䝖䜴䜵䜰䜢㉳ື䛧䜎䛩1b䝩䞊䝮⏬㠃䛷 Open 䜢䜽䝸䝑䜽䛧䜎䛩䚹

1cOpen 䝎䜲䜰䝻䜾䝪䝑䜽䝇䛷䚸experiments 䝣䜷䝹䝎䜢㑅ᢥ䛧䜎䛩䠄ึᮇタᐃ䛾ሙྜ䠅ὀグ䠖䜒䛧௵ព䛾ሙᡤ䛻䝣䜯䜲䝹䜢ಖᏑ䛧䛯ሙྜ䛿䚸䛭䛾ሙᡤ䜢㛤䛔䛶ୗ䛥䛔䚹1dಖᏑ䛧䛯䝣䜯䜲䝹䜢㑅ᢥ䛧䚸Open 䜢䜽䝸䝑䜽䛧䜎䛩䚹

1c

1d- 29 -

JP-886̲本文̲0707se.indd 28

15/07/08 17:33

2.

཯ᛂ䝥䝺䞊䝖䜢 StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛻䝉䝑䝖䛧䜎䛩䚹䠖ᶵჾ䛾ືస୰䚸䝃䞁䝥䝹䝤䝻䝑䜽䛿 100䉝㏆䛟䜎䛷⇕䛫䜙䜜䛶䛔䜎䛩䚹┤๓䛻ᶵჾ䜢౑⏝䛧䛶䛔䛯ሙྜ䚸䝃䞁䝥䝹䝤䝻䝑䜽䛜ᐊ䛻ୗ䛜䛳䛶䛛䜙సᴗ䜢⾜䛳䛶ୗ䛥䛔䚹㔜せ䠖཯ᛂ⣔ㄪᩚ⏝䛾ᾘ⪖ရ䜢ྲྀ䜚ᢅ䛖㝿䛻䛿ᚲ䛪䝟䜴䝎䞊䝣䝸䞊䛾ᡭ⿄䜢⿦╔䛧䛶ୗ䛥䛔䚹2aᶵჾṇ㠃䛾䝗䝻䜰䞊䜢㛤䛡䜎䛩䚹2b䝃䞁䝥䝹䝤䝻䝑䜽䛻཯ᛂ䝥䝺䞊䝖䜢䝉䝑䝖䛧䜎䛩䚹-Reaction Plate 䜢⏝䛔䛯ሙྜ䛿䚸A1 䛜ᕥୖ䛻䛺䜛䜘䛖䛻䝉䝑䝖䛧䜎䛩䚹-Tube 䜢䛤฼⏝䛾ሙྜ䛿䚸Tray 䛻䝉䝑䝖䛧䛶䚸䛭䛾䜎䜎䝤䝻䝑䜽䛻䝉䝑䝖䛧䜎䛩䚹㔜せ䟿䠖StepOne 䝅䝇䝔䝮䛿 4 ᮏ௨ୖ䛾䝏䝳䞊䝤䜢䝖䝺䜲䛻䝉䝑䝖䛧䛶䛟䛰䛥䛔䚹StepOnePlus 䝅䝇䝔䝮䛿 16 ᮏ௨ୖ䛾䝏䝳䞊䝤䜢⦪䠄A-H䠅2 ิ䜎䛯䛿ᶓ䠄3-10䠅2 ิ䛻୪䜉䛶䝖䝺䜲䛻䝉䝑䝖䛧䛶䛟䛰䛥䛔䚹2c䝗䝻䜰䞊䛻ᡭ䜢䛿䛥䜎䛺䛔䜘䛖䛻ὀព䛧䛶㛢䛨䜎䛩䚹- 30 -

JP-886̲本文̲0707se.indd 29

15/07/08 17:33

3.

䝷䞁䜢㛤ጞ䛧䜎䛩䚹
3a

StepOne 䝋䝣䝖䜴䜵䜰ୖ䛷 START RUN 䜢䜽䝸䝑䜽䛧䜎䛩䚹㔜せ䠖START RUN 䜢 2 ᅇ௨ୖ㐃⥆䛧䛶䜽䝸䝑䜽䛧䛺䛔䜘䛖䛻䛤ὀព䛟䛰䛥䛔䚹䜽䝸䝑䜽䛩䜛䛸䝷䞁⤊஢ᚋ䛻䝣䜯䜲䝹䛜㛤䛛䛺䛟䛺䜛➼䛾䝖䝷䝤䝹䛜Ⓨ⏕䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹

Setup ⏬㠃䜢㑅ᢥ䛧䛶䛔䜛䛸䛝

RUN ⏬㠃䜢㑅ᢥ䛧䛶䛔䜛䛸䛝ὀグ䠖䝷䞁୰䛻䝯䞁䝔䝘䞁䝇䜢⾜䛳䛯䜚䚸䝞䝑䜽䜾䝷䞁䝗䛷䜴䜱䝹䝇ᑐ⟇䝋䝣䝖䜴䜵䜰䛜㉳ື䛧䛺䛔䜘䛖䛻䛧䛶䛟䛰䛥䛔䚹䝷䞁୰䛻䛣䜜䜙䛾సᴗ䜢⾜䛖䛸䚸ṇᖖ䛺䝕䞊䝍ᅇ཰䛜⾜䜟䜜䛺䛟䛺䜛༴㝤ᛶ䛜䛒䜚䜎䛩䚹- 31 -

JP-886̲本文̲0707se.indd 30

15/07/08 17:33

4.

䝷䞁୰䛾䝰䝙䝍䝸䞁䜾⏬㠃ᕥഃ䛾 Experiment Menu 䜢㐺ᐅ㑅ᢥ䛧䛶䚸䝷䞁୰䛻䝃䜲䜽䝹䝇䝔䞊䝆䛾☜ㄆ䜔䚸⺯ග䝕䞊䝍䜢䝰䝙䝍䝸䞁䜾䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹䜎䛯ᐇ㦂᮲௳䛾タᐃ䜢⾜䛖䛣䛸䜒䛷䛝䜎䛩䚹
4a

Setup 䜢䜽䝸䝑䜽䛩䜜䜀䚸䝷䞁୰䛷䜒ᐇ㦂᮲௳䛾タᐃ䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹

4b

Run 䜢୰Ṇ䛩䜛䛸䛝䛻䛿 STOP RUN 䜢䜽䝸䝑䜽䛧䜎䛩䚹Stop Run 䝎䜲䜰䝻䜾䛜⾲♧䛥䜜䜛䛾䛷䚸௨ୗ䛾୰䛛䜙㑅ᢥ䛧䜎䛩䚹- Stop Immediately : ༶ᗙ䛻䝷䞁䜢୰Ṇ䛧䜎䛩䚹- Stop after Current Cycle / Hold : ᭱ᚋ䛾 Cycle 䝇䝔䞊䝆䜎䛯䛿 Hold 䛜⤊஢䛧䛶䛛䜙䝷䞁䜢୰Ṇ䛧䜎䛩䚹- Cancel : 䝷䞁䛾୰Ṇ䜢䜻䝱䞁䝉䝹䛧䚸䝷䞁䜢⥅⥆䛧䛶⾜䛔䜎䛩䚹4cAmplification Plot : 䝷䞁୰䛻⺯ගᙉᗘ䛾ቑᖜ᭤⥺䜢☜ㄆ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹4d

Temperature Plot : Sample ⁐ᾮ䛸 Block ཬ䜃 Heat Cover 䛾

4e

Run Method : タᐃ䛧䛯 Run Method 䜢☜ㄆ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹䜎䛯䝷䞁୰䛻䝃䜲䜽䝹ᩘ䛻ኚ᭦䛧䛯䜚䚸ᗘ䜢䝰䝙䝍䝸䞁䜾䛷䛝䜎䛩䚹

Melt Curve 䛾䝥䝻䜾䝷䝮䜔䝩䞊䝹䝗䜢㏣ຍ䛩䜛䛣䛸䜒䛷䛝䜎䛩䚹ὀグ䠖䝷䞁㛤ጞ᫬䛻 Melt Curve 䛾 Stage 䛜ධ䛳䛶䛔䜛䛸䚸䝷䞁୰䛻䝥䝻䜾䝷䝮ኚ᭦䛩䜛䛣䛸䛿䛷䛝䜎䛫䜣䚹

4f

Notification Setting : 䜶䝷䞊䛜Ⓨ⏕䛧䛯᫬䛾⮬ື䝯䞊䝹㐃⤡ᶵ⬟䜢タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹ὀグ䠖䛣䛾ᶵ⬟䜢฼⏝䛩䜛䛯䜑䛻䛿䝛䝑䝖䝽䞊䜽䛻᥋⥆䛧䛶䛔䜛䛣䛸䛜ᚲせ䛷䛩䚹- 32 -

JP-886̲本文̲0707se.indd 31

15/07/08 17:33

5.

ᐇ㦂䛜⤊஢䛧䛯䜙䚸཯ᛂ䝥䝺䞊䝖䜢 StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛛䜙ྲྀ䜚ฟ䛧䜎䛩䚹䠖ᶵჾ䛾ືస୰䚸䝃䞁䝥䝹䝤䝻䝑䜽䛿 100䉝㏆䛟䜎䛷⇕䛫䜙䜜䛶䛔䜎䛩䚹┤๓䛻ᶵჾ䜢౑⏝䛧䛶䛔䛯ሙྜ䚸䝃䞁䝥䝹䝤䝻䝑䜽䛜ᐊ

6.

䛻ୗ䛜䛳䛶䛛䜙సᴗ䜢⾜䛳䛶ୗ䛥䛔䚹5aᶵჾṇ㠃䛾䝗䝻䜰䞊䜢㛤䛝䜎䛩䚹5b཯ᛂ䝥䝺䞊䝖䜢䝃䞁䝥䝹䝤䝻䝑䜽䛛䜙ྲྀ䜚ฟ䛧䜎䛩䚹5c䝗䝻䜰䞊䛻ᡭ䜢䛿䛥䜎䛺䛔䜘䛖䛻ὀព䛧䛶㛢䛨䜎䛩䚹䠄䜸䝥䝅䝵䞁䠅StepOne 䛚䜘䜃 StepOnePlus 䝅䝇䝔䝮䛾䝅䝱䝑䝖䝎䜴䞁㐃⥆䛧䛶䝷䞁䜢⾜䜟䛺䛔ሙྜ䛿䚸䝷䞁⤊஢᫬䛻 StepOne 䛚䜘䜃 StepOnePlus 䛾㟁※䜢ษ䛳䛶䛔䛯䛰䛟䛣䛸䛜䛷䛝䜎䛩䚹䝕䞊䝍ゎᯒ䛿䝋䝣䝖䜴䜵䜰༢⊂䛷⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹䜎䛯 StepOne 䛚䜘䜃 StepOnePlus 䝍䝑䝏䝟䝛䝹ᕥୗ䛾䝪䝍䞁䜢䜽䝸䝑䜽䛩䜛䛸䚸䝇䝍䞁䝞䜲䝰䞊䝗䛻䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

- 33 -

JP-886̲本文̲0707se.indd 32

15/07/08 17:33

Standard Curve Method 䝕䞊䝍䛾ゎᯒ䞉ฟຊ䝷䞁䛜⤊஢䛧䛯䜙ゎᯒ䜢⾜䛔䜎䛩䚹1.

䝣䜯䜲䝹ྑୖ䛾 Analyze 䜢䜽䝸䝑䜽䛩䜛䛸䝕䞊䝍ゎᯒ䛜⾜䜟䜜䜎䛩䚹ὀグ䠖ึᮇタᐃ䛷䛿䝋䝣䝖䜴䜵䜰䛻䜘䜛⮬ືゎᯒ䛜⾜䜟䜜䜎䛩䚹2.

ゎᯒ䛜⤊஢䛩䜛䛸䚸Experiment Menu ෆ Analysis>Amplification Plot 䛜⾲♧䛥䜜䜎䛩䚹䛣䛾≧ែ䛷䚸඲䛶䛾ゎᯒ䛜⾜䜟䜜䚸ᐃ㔞⤖ᯝ䜎䛷⟬ฟ䛥䜜䛶䛔䜎䛩䚹

Plot 䝟䝛䝹
View 䝍䝤

2a⏬㠃ᕥ䛾 Experiment Menu 䛾 Analysis 䛾㡯┠䜢㑅ᢥ䛧䛶䚸Plot 䝟䝛䝹䛻⾲♧䛥䜜䜛⏬㠃䜢ኚ᭦䛷䛝䜎䛩䚹䜎䛯ᚲせ䛻ᛂ䛨䛶 Setup 䛛䜙ᐇ㦂᮲௳䛾タᐃ䞉ኚ᭦䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹2b
3.

View 䝍䝤䜢䜽䝸䝑䜽䛧䛶䚸Plate Layout 䛸 Well Table 䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

Amplification Plot 䛷ゎᯒタᐃ䜢☜ㄆ䛧䜎䛩䚹
Target 䛤䛸䛻 Baseline 䛾☜ㄆ䛸 Threshold Line 䛾☜ㄆ䜢⾜䛔䜎䛩䚹- 34 -

JP-886̲本文̲0707se.indd 33

15/07/08 17:33

ゎᯒタᐃ䛾䜺䜲䝗䝷䜲䞁
StepOne 䝋䝣䝖䜴䜵䜰䛿ୗᅗ䛾䜘䛖䛺඾ᆺⓗ䛺 Amplification Plot 䛾䝕䞊䝍䛜䛒䜛䛸䛝䛻䚸⮬ືⓗ䛻 Baseline䛸 Threshold Line 䜢タᐃ䛧䜎䛩䚹a : 䝥䝷䝖䞊㡿ᇦb : ୍ḟ㛵ᩘⓗቑᖜ㡿ᇦc : ᣦᩘ㛵ᩘⓗቑᖜ㡿ᇦ䠄ᗄఱᏛ఩┦䠅d : 䝞䝑䜽䜾䝷䜴䞁䝗e : 䝧䞊䝇䝷䜲䞁㔜せ䠖䝤䝻䝑䜽䛾ởᰁ䜔䝁䞁䝍䝭䞉䝢䝨䝑䝔䜱䞁䜾䜶䝷䞊䛺䛹䛻䜘䜚䚸Amplification Plot 䛾Ἴᙧ䛜ኚ䜟䜛䛸䚸
StepOne 䝋䝣䝖䜴䜵䜰䛿ṇ☜䛺 Baseline(p-31)䛸 Threshold Line(p-32)䜢タᐃ䛩䜛䛣䛸䛜䛷䛝䛺䛟䛺䜛䛣䛸䛜䛒䜚䜎䛩䚹䜰䝥䝷䜲䝗䝞䜲䜸䝅䝇䝔䝮䝈䛷䛿䚸ゎᯒᚋ Amplification Plot ⏬㠃䛷 Baseline 䛸 Threshold Line 䜢☜ㄆ䛩䜛䛣䛸䜢᥎ዡ䛧䜎䛩䚹㢧ⴭ䛻䜲䝺䜼䝳䝷䞊䛺䝕䞊䝍䛾䜴䜵䝹䛜䛒䜛᫬䛻䛿䚸䛭䜜䜙䛾䜴䜵䝹䜢ゎᯒᑐ㇟እ䛻タᐃ䠄Omit䠅䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹43 䝨䞊䝆䜢䛤ཧ↷䛟䛰䛥䛔䚹- 35 -

JP-886̲本文̲0707se.indd 34

15/07/08 17:33

3aBaseline 䛾☜ㄆ
3aGraph Type 䛜 Liner 䛷䛒䜛䛣䛸䜢☜ㄆ䛧䜎䛩䚹

3bTarget 䜢㐺ᐅタᐃ䛧䜎䛩䚹

3cshow:䛾 Baseline 䛾䝏䜵䝑䜽䝪䝑䜽䝇䛻䝏䜵䝑䜽䜢ධ䜜䜎䛩䚹Baseline 䛿⺯ගᙉᗘ䛜 0 䛾⠊ᅖ䛾タᐃ䛷䛩䚹Start 䛛䜙 Stop䜎䛷䚸⺯ගᙉᗘ䛜 0 䛾㡿ᇦ䜢ṇ䛧䛟タᐃ䛷䛝䛶䛔䜛䛛䛹䛖䛛☜3cㄆ䛧䜎䛩䚹ྑୖᅗ䛾䜘䛖䛺ቑᖜ᭤⥺䛷䛒䜜䜀䚸䛭䛾䜎䜎ḟ䛾䝇3b䝔䝑䝥䛻㐍䜏䜎䛩䚹䜒䛧ྑ୰ኸᅗ䛾䜘䛖䛻䚸ᣦᩘ㛵ᩘⓗ䛻ቑᖜ䛧䛶䛔䛺䛔㡿ᇦ䜢Baseline 䛾⠊ᅖ䛸䛧䛶ᣦᐃ䛧䛶䛔䛺䛔ሙྜ䛿䚸Manual 䛷Baseline タᐃ䜢⾜䛖䛣䛸䛜ᚲせ䛷䛩䚹
Manual 䛷䛾 Baseline タᐃ䛿 Target ẖ䛻䚸඲䛶䛾䝃䞁䝥䝹䛜ቑᖜ䛧䛶䛔䛺䛔䝃䜲䜽䝹䛾⠊ᅖ䜢ᣦᐃ䛩䜛ᙧ䛷⾜䛔䜎䛩䚹Plot 䝟䝛䝹ୗẁ䛾 Options 䝍䝤ෆ䛷䚸Threshold Auto 䛸䚸Auto
Baseline 䛾䝏䜵䝑䜽䜢 2 䛴䛸䜒䛻እ䛧䜎䛩䚹䛩䜛䛸 Amplification
Plot ᶓ㍈䛾୍␒ᕥഃ䛻㻌 㻌 㻌 䝬䞊䜽䛜⾲♧䛥䜜䜎䛩䚹䛣䛾䝬䞊䜽䛿䝬䜴䝇䛾䝫䜲䞁䝍䜢ྜ䜟䛫䛶ᶓ䛻䝗䝷䝑䜾䛧䛶ື䛛䛩䛣䛸䛜䛷䛝䜎䛩䛾䛷䚸඲䛶䛾䝃䞁䝥䝹䛜ቑᖜ䛧䛶䛔䛺䛔⠊ᅖ䛾ᕥ➃䛻⥳䛾䚸ྑ➃䛻㉥䛾

䜢䚸䛭䜜䛮䜜ྜ䜟䛫䜎䛩䚹- 36 -

JP-886̲本文̲0707se.indd 35

15/07/08 17:33

3d

Threshold Line 䛾☜ㄆ
3dGraph Type 䜢 Liner 䛛䜙 Log 䛻ኚ᭦䛧䜎䛩䚹

3eTarget 䜢㐺ᐅタᐃ䛧䜎䛩䚹

3f

show:䛾 Threshold 䛾䝏䜵䝑䜽䝪䝑䜽䝇䛻䝏䜵䝑䜽䜢ධ䜜䜎䛩䚹

Threshold Line 䛿䚸ᣦᩘ㛵ᩘⓗ䛻ቑᖜ䛧䛶䛔䜛㡿ᇦ䛻タᐃ䛥䜜䛶䛔䜛䛣䛸䛜ᚲせ䛷䛩䚹⦪㍈䜢ᑐᩘ㍈䛻䛩䜛䛸䚸ᣦᩘ㛵ᩘⓗ䛻ቑᖜ䛧䛶䛔䜛㡿ᇦ䛜┤⥺䛷⾲♧䛥䜜䜎䛩䚹඲䛶䛾ቑᖜ䝥䝻䝑䝖䛜┤⥺䛷⾲♧䛥䜜䛶䛔䜛㡿ᇦ䛻 Threshold Line 䛜タᐃ䛥䜜䛶䛔䜛䛣䛸䜢䛤☜ㄆୗ䛥䛔䚹3f3e඲䛶䛾ቑᖜ䝥䝻䝑䝖䛜┤⥺䛷⾲♧䛥䜜䛶䛔䜛㡿ᇦ䛻 ThresholdLine 䛜タᐃ䛥䜜䛶䛔䛺䛔ሙྜ䜔䚸Manual 䛷 Baseline 䜢タᐃ䛧䛯䛸䛝䛿䚸Manual 䛷 Threshold Line 䜢タᐃ䛩䜛䛣䛸䛜ᚲせ䛷䛩䚹Plot 䝟䝛䝹⏬㠃ୗ䛾 Options 䝍䝤ෆ䚸Threshold Auto 䛾䝏䜵䝑䜽䝪䝑䜽䝇䜢እ䛩䛸䚸Amplification Plot ⏬㠃ୖ䛷 ThresholdLine 䛜ୖୗ䛻ື䛛䛫䜛䜘䛖䛻䛺䜚䜎䛩䚹඲䛶䛾䝃䞁䝥䝹䛜ྑୖ䛜䜚䛾┤⥺䛷ቑᖜ䛧䛶䛔䜛㡿ᇦ䛻 Threshold Line 䜢タᐃ䛧䜎䛩䚹䜎䛯䚸Options 䝍䝤䛷 Threshold 䛾್䜢┤᥋ධຊ䛩䜛䛣䛸䜒䛷䛝䜎䛩䚹

- 37 -

JP-886̲本文̲0707se.indd 36

15/07/08 17:33

4.

ゎᯒ䛾タᐃ䛾☜ㄆ䛜⤊஢䛧䛯䜙䚸Standard Curve 䜢☜ㄆ䛧䜎䛩䚹

4aExperiment Menu 䛷 Standard Curve 䜢䜽䝸䝑䜽䛧䜎䛩䚹

4b

Target 䛿 All 䜢㑅ᢥ䛧䜎䛩䚹4cPlot Color 䛿 Default 䜢㑅ᢥ䛧䜎䛩䚹

4dShow/Hide the plot legend 䜢䜽䝸䝑䜽䛧䜎䛩䚹

4eTarget ẖ䛾 Slope 䛸 R2 䛸 Eff%䜢☜ㄆ䛧䜎䛩䚹
-Slope : ᳨㔞⥺䛾ഴ䛝䛷䛩䚹PCR ຠ⋡䠄Eff%䠅䜢཯ᫎ䛧䚸ຠ⋡䛜 100%䛾᫬䛻䛿-3.32 䛻䛺䜚䜎䛩䚹- R2 : ┦㛵ಀᩘ䛷䛩䚹CT ್䛸ึᮇ㔞䛜᏶඲䛻┦㛵䛧䛶䛔䜛䛸䛝䛻䛿 1.00 䛻䛺䜚䜎䛩䚹R2 ್䛿 0.990 ௨ୖ䛒䜛䛣䛸䛜ᮃ䜎䛧䛔䛷䛩䚹- Eff% : PCR 䛾ቑᖜຠ⋡䜢 Slope ್䛛䜙⟬ฟ䛧䛯ᩘᏐ䛷䛩䚹

4b

4c

4d
4a

4e

- 38 -

JP-886̲本文̲0707se.indd 37

15/07/08 17:33

5.

Melt Curve ⏬㠃䠄SYBR Green 䜰䝑䝉䜲䛾ሙྜ䛾䜏䠅Melt Curve ⏬㠃䛷஧ḟ⏘≀䛾᭷↓䜢☜ㄆ䛧䜎䛩䚹

5aExperiment Menu 䛷 Melt Curve 䜢䜽䝸䝑䜽䛧䜎䛩䚹5bPlot 䛜 Derivative Reporter 䛻䛺䛳䛶䛔䜛䛣䛸䜢☜ㄆ䛧䜎䛩䚹

5cTarget 䜢㑅ᢥ䛧䜎䛩䚹5d䝢䞊䜽䛜༢⊂䛛」ᩘ䛛☜ㄆ䛧䜎䛩䚹ୗᅗ䛾㏻䜚༢⊂䛷䛒䜜䜀ၥ㢟䛒䜚䜎䛫䜣䚹䜒䛧」ᩘ䛾䝢䞊䜽䛜䛒䛳䛯ሙྜ䛿䚸ᐇ㦂⣔䛾᭱㐺໬䜢⾜䛳䛶䛔䛯䛰䛟䛣䛸䛜ᚲせ䛷䛩䚹

5eTarget 䛜」ᩘ䛒䜛䛸䛝䛻䛿 1 䛴䛪䛴ษ䜚᭰䛘䛶☜ㄆ䛧䜎䛩䚹

5b

5c

5a

5d

- 39 -

JP-886̲本文̲0707se.indd 38

15/07/08 17:33

6.

View Well Table 䝍䝤⏬㠃䛷 CT ್䜢☜ㄆ䛧䜎䛩䚹
Well, Omit䠄ゎᯒ䛛䜙๐㝖䛧䛯䛔㝿䛻䝏䜵䝑䜽䛧䜎䛩䠅, Flag, Sample Name, Target Name, Task, Dyes, CT䠄ቑᖜ᭤⥺䛜 Threshold Line 䛸஺ᕪ䛧䛯䝃䜲䜽䝹ᩘ䠅, CT Mean䠄CT 䛾ᖹᆒ್䠅, CT SD䠄CT 䛾 SD䠅, Rn䠄䝧䞊䝇䝷䜲䞁⿵ṇ๓䛾཯ᛂ⤊஢᫬䛾䝅䜾䝘䝹್䠅, ǻRn䠄䝧䞊䝇䝷䜲䞁⿵ṇᚋ䛾཯ᛂ⤊஢᫬䛾䝅䜾䝘䝹್䠅,
Quantity䠄ྛ䜴䜵䝹䛾ᐃ㔞⤖ᯝ䠅, Quantity Mean䠄ᐃ㔞⤖ᯝ䛾ᖹᆒ್䠅, Quantity SD䠄ᐃ㔞⤖ᯝ䛾ᖹᆒ್
SD䠅䛺䛹䛜⾲♧䛥䜜䜎䛩䚹
6aView Well Table 䜢䜽䝸䝑䜽䛧䜎䛩䚹

6bGroup By 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙 Replicate 䜢㑅ᢥ䛧䜎䛩䚹

6cCT 䜹䝷䝮䛾್䜢☜ㄆ䛧䜎䛩䚹䛷䛝䜜䜀 8< CT <35 䛾⠊ᅖ䛻ධ䜛䛣䛸䛜ᮃ䜎䛧䛔䛷䛩䚹CT ್䛿 Threshold Line 䜎䛷⺯ගᙉᗘ䛜㐩䛧䛯䝃䜲䜽䝹ᩘ䛷䛩䚹8 䝃䜲䜽䝹ᮍ‶䛾᫬䛿䚸཯ᛂᾮ୰䛾䝔䞁䝥䝺䞊䝖㔞䛜ከ䛩䛞䜎䛩䚹35 䝃䜲䜽䝹䜢㉸䛘䜛᫬䛻䛿䚸䝍䞊䝀䝑䝖ศᏊ䛾ึᮇ㔞䛜㠀ᖖ䛻ᑡ䛺䛔䛷䛩䚹CT ್䛜 35 䝃䜲䜽䝹䜢㉸䛘䜛䛸ᶆ‽೫ᕪ䜒኱䛝䛟䛺䜚䜎䛩䚹
6a
6b

7.

6c

඲䛶䛾☜ㄆ䜢⾜䛳䛯䜙 View Well Table 䝍䝤⏬㠃䛷ᐃ㔞್䜢☜ㄆ䛧䜎䛩䚹㻌 㻌 㻌 - Group By 䛾䝥䝹䝎䜴䞁䝯䝙䝳䞊䛷⾲♧䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹㻌 㻌 㻌 - 䜲䝺䜼䝳䝷䞊䛺䜴䜵䝹䛜䛒䜛䛸䛝䛻䛿䚸Omit 䛾䝏䜵䝑䜽䜢ධ䜜䚸⏬㠃ྑୖ䛾 Analyze 䜢䜽䝸䝑䜽䛩䜜䜀㻌 㻌ゎᯒᑐ㇟እ䛻䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹7a⏬㠃ᕥ䛾 Experiment Menu 䛾 Analysis 䛾㡯┠䜢㑅ᢥ䛧䛶䚸Plot 䝟䝛䝹䛻⾲♧䛥䜜䜛⏬㠃䜢ኚ᭦䛷䛝䜎
䛩䚹

7b

8.

䜎䛯ᚲせ䛻ᛂ䛨䛶 Setup 䛛䜙ᐇ㦂᮲௳䛾タᐃ䞉ኚ᭦䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹ゎᯒ䛜᏶஢䛧䛯䜙⏬㠃ᕥୖ Save 䜢䜽䝸䝑䜽䛧䛶䚸ゎᯒタᐃ䜢ಖᏑ䛧䜎䛩䚹ゎᯒ䛜᏶஢䛧䛯䜙⏬㠃ᕥୖ Save 䜢䜽䝸䝑䜽䛧䛶䚸ゎᯒタᐃ䜢ಖᏑ䛧䜎䛩䚹- 40 -

JP-886̲本文̲0707se.indd 39

15/07/08 17:33

9.

ゎᯒ⤖ᯝ䜔䜾䝷䝣䜢䝣䜯䜲䝹䛷ฟຊ䛷䛝䜎䛩䚹- ⏬㠃ྑୖ䛾 Export 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙 Export 䜢㑅ᢥ䛩䜛䛸䚸View Well Table 䝍䝤⏬㠃䛾᝟ሗ䜢ึ䜑䚸඲䛶䛾䝕䞊䝍䜢⾲ᙧᘧ䛷ฟຊ䛷䛝䜎䛩䚹䛣䛾䛸䛝䚸ฟຊ䝣䜷䞊䝬䝑䝖䜢 Excel 䜎䛯䛿 Text ᙧᘧ䛛䜙㑅ᢥ䛷䛝䜎䛩䚹- ⏬㠃ྑୖ䛾 Export 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙 Send to Slides 䜢㑅ᢥ䛩䜛䛸䚸PowerPoint ᙧᘧ䛷⏬ീ䜢ฟຊ䛷䛝䜎䛩䚹
ὀグ䠖䛣䛾ᶵ⬟䛿 PowerPoint 䛜䜲䞁䝇䝖䞊䝹䛥䜜䛶䛔䜛 PC ୖ䛾䜏䛷᭷ຠ䛷䛩䚹ὀグ䠖䛣䛾ᶵ⬟䜢฼⏝䛩䜛๓䛻䚸඲䛶䛾 PowerPoint 䛾䜰䝥䝸䜿䞊䝅䝵䞁䛜⤊஢䛧䛶䛔䜛䛣䛸䜢䛤☜ㄆୗ䛥䛔䚹
PowerPoint 䛾䜰䝥䝸䜿䞊䝅䝵䞁䛜㉳ື䛧䛶䛔䜛䛸䚸⏬ീ䛜ฟຊ䛥䜜䛺䛔䜎䜎䚸PowerPoint 䛾䜰䝥䝸䜿䞊䝅䝵䞁䜒⤊஢䛧䜎䛩䚹䛣䛾᫬㛤䛔䛶䛔䛯䝣䜯䜲䝹䛜䛒䜛䛸䚸᭦᪂᝟ሗ䛿ಖᏑ䛥䜜䛪䛻ᙉไ⤊஢䛧䜎䛩䚹- Plot 䝟䝛䝹⏬㠃䛷ྑ䜽䝸䝑䜽>Save As 䜢㑅ᢥ䛩䜛䛸䚸⏬ീ䜢 JPEG ᙧᘧ䛷ಖᏑ䛷䛝䜎䛩䚹-඲䛶䛾⏬ീཬ䜃⾲䜢༳ๅ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹䝥䝸䞁䝍䞊䝗䝷䜲䝞䞊䛻 PDF 䝣䜯䜲䝹సᡂ⏝䛾䝗䝷䜲䝞䞊䠄CutePDF Writer䠅䛜䜲䞁䝇䝖䞊䝹䛥䜜䛶䛔䜎䛩
䛾䛷䚸PDF 䝣䜯䜲䝹䛸䛧䛶ಖᏑ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹- 41 -

JP-886̲本文̲0707se.indd 40

15/07/08 17:33

㏣ຍ☜ㄆ஦㡯䛣䛣䛷䛿䝕䞊䝍ゎᯒ᫬䛻᭷⏝䛺᝟ሗ䛻䛴䛔䛶䛤ㄝ᫂䛧䜎䛩䚹

QC Summary ⏬㠃Analyze 䜢⾜䛳䛯ᚋ䛻䚸View Plate Layout ⏬㠃䛾䜴䜵䝹ୖ䛻䝣䝷䜾䠄㯤Ⰽ䛾୕ゅ䝬䞊䜽䠅䛜⾲♧䛥䜜䜛䛣䛸䛜䛒䜚䜎䛩䚹䝣䝷䜾䛿⾲♧䛥䜜䛯䜴䜵䝹䛻ఱ䛛䛧䜙䛾ၥ㢟䛜䛒䜛䛣䛸䜢ᥦ♧䛧䜎䛩䚹䝣䝷䜾ෆ䛾ᩘᏐ䛿☜ㄆ䛥䜜䛯ၥ㢟䛾ᩘ䜢♧䛧䜎䛩䚹
QC Summary ⏬㠃䛷䛿䝣䝷䜾䛾ලయⓗ䛺ෆᐜ䚸ཬ䜃䛭䛾ᑐ⟇᪉ἲ䜢☜ㄆ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

1.

Experiment Menu 䛷 QC Summary 䜢䜽䝸䝑䜽䛧䜎䛩䚹2.

䛂Flag Summary䛃䜢☜ㄆ䛧䜎䛩䚹

3.

䛂Flag Details䛃䝔䞊䝤䝹䛾䛂Frequency䛃䛚䜘䜃䛂Wells䛃䜹䝷䝮䜢ㄪ䜉䚸ၥ㢟䛾ෆᐜ䛸䜴䜵䝹ᩘ䚸䛚䜘䜃䜴䜵䝹䛾఩⨨䜢☜ㄆ䛧䜎䛩䚹

4.

䝣䝷䜾䛾⾜䜢䜽䝸䝑䜽䛩䜛䛸䚸ୗẁ䛻䛭䛾䝣䝷䜾䛾ヲ⣽䛜⾲♧䛥䜜䜎䛩䚹

2

3
1
4

- 42 -

JP-886̲本文̲0707se.indd 41

15/07/08 17:33

Standard Curve Method ᐇ㦂䛷䛿䚸ᐇ㦂䝕䞊䝍䛾≧ἣ䛻䜘䜚ୗグ䛾䝣䝷䜾䛜⾲♧䛥䜜䜎䛩䚹
䝣䝷䜾
AMPNC
BADROXBLFAILCTFAILEXPFAILHIGHSD
MTP
NOAMP
NOISE
NOSIGNAL
OFFSCALEOUTLIERRG
SPIKE
THOLDFAILෆᐜ䝛䜺䝔䜱䝤䝁䞁䝖䝻䞊䝹䛷ቑᖜ䝟䝑䝅䝤䝸䝣䜯䝺䞁䝇್䛻␗ᖖ䛒䜚䝧䞊䝇䝷䜲䞁⮬ືタᐃ᫬䛻ၥ㢟☜ㄆCT ್⮬ື⟬ฟ᫬䛻ၥ㢟☜ㄆᣦᩘ㛵ᩘⓗቑᖜ㡿ᇦ䛜☜ㄆ䛥䜜䛺䛔䝸䝥䝸䜿䞊䝖䜾䝹䞊䝥ෆ䛾ᶆ‽೫ᕪ䛜㧗䛔Tm 䝢䞊䜽䛜」ᩘ䛒䜛ὀ䠖䛣䛾䝣䝷䜾䛿 Melt Curve 䛾䝕䞊䝍䛜ྵ䜎䜜䛶䛔䜛ሙྜ䛾䜏⾲♧䛥䜜䜎䛩ቑᖜ䛺䛧௚䛾䜴䜵䝹䛸ẚ㍑䛧䛶䝜䜲䝈䛜㧗䛔⺯ග್䛜᳨ฟ䛥䜜䛺䛔⺯ග್䛜䜸䝣䝇䜿䞊䝹䝸䝥䝸䜿䞊䝖䜾䝹䞊䝥ෆ䛻␗ᖖ್䛜䛒䜛䝇䝟䜲䜽䝜䜲䝈᭷䜚
Threshold ⮬ືタᐃ᫬䛻ၥ㢟☜ㄆ

Omit Well
௵ព䛾䜴䜵䝹䜢ゎᯒᑐ㇟እ䛻ᣦᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹
๓㏙䛾 View Well Table 䝍䝤⏬㠃䛷ᑐ㇟እ䛻ᣦᐃ䛩䜛᪉ἲ䠄40 䝨䞊䝆㻌 䝇䝔䝑䝥 7䠅䛾䜋䛛䚸View Plate
Layout 䝍䝤⏬㠃䛷䜒ゎᯒᑐ㇟እ䛻ᣦᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

1.

View Well Table 䝍䝤⏬㠃ୖ䛷䚸ゎᯒᑐ㇟እ䛻䛩䜛௵ព䛾䜴䜵䝹䜢㑅ᢥ䛧䜎䛩䚹

2.

䛭䛾䜎䜎ྑ䜽䝸䝑䜽䛩䜛䛸䝯䝙䝳䞊䛜⾲♧䛥䜜䜛䛾䛷䚸Omit 䜢㑅ᢥ䛧䜎䛩䚹

3.

ゎᯒᑐ㇟እ䛻ᣦᐃ䛥䜜䛯䜴䜵䝹䛾ᕥୖ䛻㉥䛔㽢䝬䞊䜽䛜⾲♧䛥䜜䜎䛩䚹

4.

Reanalyze 䝪䝍䞁䜢䜽䝸䝑䜽䛩䜛䛸䚸ᣦᐃ䛧䛯䜴䜵䝹䛾䝕䞊䝍䜢㝖እ䛧䛯ゎᯒ䛜⾜䜟䜜䜎䛩䚹

5.

ゎᯒᑐ㇟䛻ᡠ䛩䛸䛝䛿䚸䜴䜵䝹䜢㑅ᢥ䛧䛶ྑ䜽䝸䝑䜽䝯䝙䝳䞊䛛䜙 Include䜢㑅ᢥ䛧䜎䛩䚹

- 43 -

JP-886̲本文̲0707se.indd 42

15/07/08 17:33

Baseline 䛾ᡭືタᐃ䠖௵ព䛾䜴䜵䝹䛾䜏ᣦᐃ䛧䛯䜴䜵䝹䛾 Baseline 䜢ᡭື䛷タᐃ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹⮬ືゎᯒ䜢⾜䛳䛯䛸䛝䛻䚸Ṥ䛹䛾䜴䜵䝹䛿ၥ㢟䛺䛔䛾䛻䚸䛤䛟୍㒊䛾䜴䜵䝹䛾䜏 Baseline 䛾タᐃ䛜Ⰻዲ䛻⾜䜟䜜䛺䛛䛳䛯䛸䛝䛻䚸௵ព䛾䜴䜵䝹䛾䜏ᡭື䛷Baseline 䜢タᐃ䛧䚸෌ゎᯒ䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹1.

䛒䜙䛛䛨䜑 Amplification Plot ⏬㠃䛷䚸Baseline 䜢ኚ᭦䛩䜛䜴䜵䝹䛸䚸タᐃ䛩䜛 Start 䛸 Stop 䛾ᩘ್䜢☜ㄆ䛧䚸᥍䛘䛶䛚䛝䜎䛩䚹2.

⏬㠃ྑୖ䛾 Analysis Settings 䜢䜽䝸䝑䜽䚸䜎䛯䛿 Analysis 䝯䝙䝳䞊䛛䜙 Analysis Settings 䜢㑅ᢥ䛩
䜛䛸䚸Analysis Settings ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹

3.

Advanced Settings 䝍䝤䜢䜽䝸䝑䜽䛧䜎䛩䚹4.

ྛ䜴䜵䝹䛾 Baseline 䛾タᐃ䝸䝇䝖䛜⾲♧䛥䜜䜛䛾䛷䚸ᡭື䛷 Baseline 䜢タᐃ䛩䜛௵ព䛾䜴䜵䝹䜢䜽䝸䝑䜽䛧䛶䝝䜲䝷䜲䝖䛷㑅ᢥ䛧䜎䛩䠄ྠ᫬䛻」ᩘ䛾䜴䜵䝹䜢㑅ᢥ䛩䜛䛣䛸䜒ྍ⬟䛷䛩䠅䚹

5.

⏬㠃ྑഃ䛾[Use CT Settings Defined for Target]ཬ䜃[Automatic Baseline]䛾䝏䜵䝑䜽䜢እ䛧䜎䛩䚹

6.

Baseline Start Cycle:䛸 End Cycle:䛾㡯┠䛜ධຊ䛷䛝䜛䜘䛖䛻䛺䜛䛾䛷䚸㐺ᐅ௵ព䛾್䜢ධຊ䛧䜎䛩䚹

7.

⏬㠃ୗ䛾 Apply Analysis Settings 䜢䜽䝸䝑䜽䛩䜛䛸䚸タᐃ䛧䛯ᡭື䛾 Baseline 䜢཯ᫎ䛧䛯෌ゎᯒ䛜⾜
䜟䜜䜎䛩䚹
3

5

6

4

7

- 44 -

JP-886̲本文̲0707se.indd 43

15/07/08 17:33

Relative Standard Curve ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ
Advanced Setup 䛛䜙 Relative Standard Curve 䜢㑅ᢥ䛧䛯᫬䛾タᐃ᪉ἲ䛻䛴䛔䛶䛤ㄝ᫂䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Standard Curve 䛾タᐃ䛸␗䛺䜛㡯┠䛾䜏䛤ㄝ᫂⮴䛧䜎䛩䚹䛭䛾௚䛾㡯┠䛾タᐃ᪉ἲ䛻䛴䛝䜎䛧䛶䛿䚸Standard Curve䛷䛾ヱᙜ㡯┠䜢䛤ཧ↷ୗ䛥䛔䚹

Experiment Properties ⏬㠃䠖
”What type of Experiment do you want to set up?”ෆ䛾 Quantitation – Relative Standard Curve 䜢䜽䝸䝑䜽䛧䜎䛩䚹

Plate Setup ⏬㠃䠖㻌
Define Target and Samples 䝍䝤䛷䠰䠽䡎䡃䡁䡐䛸 Sample 䛾タᐃ䜢⾜䛔䜎䛩䚹Define Biological Replicate Groups 䛿 Relative Standard Curve 䛷䛿౑⏝䛧䜎䛫䜣䛾䛷タᐃ䛿୙せ䛷䛩䚹- 45 -

JP-886̲本文̲0707se.indd 44

15/07/08 17:33

Assay Targets and Samples 䝍䝤䛷“Select relative quantitation settings.”䛾㡯┠䛜㏣ຍ䛥䜜䛶䛔䜎䛩䚹
1.

“Select relative quantitation settings.”௨እ䛾タᐃ䜢㐺ᐅ⾜䛔䜎䛩䚹

2.

Reference Sample 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙䚸┦ᑐ್䜢⟬ฟ䛩䜛䛯䜑䛾ᇶ‽䛸䛺䜛䝃䞁䝥䝹䜢㑅ᢥ䛧䜎䛩䚹䝋䝣䝖䜴䜵䜰䛿䛣䛣䛷タᐃ䛥䜜䛯䝃䞁䝥䝹䛾್䜢”1”䛸䛧䛯䛸䛝䛾┦ᑐ್䜢⟬ฟ䛧䜎䛩䚹

3.

Endogenous Control 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛾 Target 䜢㑅ᢥ䛧䜎䛩䚹䝋䝣䝖䜴䜵䜰䛿䛣䛣䛷タᐃ䛧䛯 Target 䛾ᐃ㔞್䜢⏝䛔䛶䚸䝃䞁䝥䝹ẖ䛻⮬ື⿵ṇ䜢⾜䛔䜎䛩䚹

4.

Endogenous Control 䛷 Multiple Controls 䜢㑅ᢥ䛩䜛䛸䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛸䛧䛶౑⏝䛩䜛 Target 䛾㑅ᢥ⏬
㠃䛜⾲♧䛥䜜䜎䛩䚹䛣䛣䛻䝏䜵䝑䜽䜢ධ䜜䛯 Target ⩌䛾ᖹᆒ್䛜ෆᅾᛶ䝁䞁䝖䝻䞊䝹್䛸䛧䛶⿵ṇィ⟬䛻⏝䛔䜙䜜䜎䛩䚹䝏䜵䝑䜽䜢ධ䜜䛯䜙 OK 䜢䜽䝸䝑䜽䛧䜎䛩䚹2
3
4- 46 -

JP-886̲本文̲0707se.indd 45

15/07/08 17:33

Relative Standard Curve 䝕䞊䝍ゎᯒ
Advanced Setup 䛛䜙 Relative Standard Curve 䜢㑅ᢥ䛧䛯᫬䛾䝕䞊䝍ゎᯒ䛻䛴䛔䛶䛤ㄝ᫂䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Standard Curve 䛾タᐃ䛸␗䛺䜛㡯┠䛾䜏䛤ㄝ᫂⮴䛧䜎䛩䚹䛭䛾௚䛾㡯┠䛾ゎᯒ᪉ἲ䛻䛴䛝䜎䛧䛶䛿䚸Standard Curve䛷䛾ヱᙜ㡯┠䜢䛤ཧ↷ୗ䛥䛔䚹

Relative Standard Curve 䜢㑅ᢥ䛧䛯ሙྜ䚸Analysis ⏬㠃䛻 Gene Expression ⏬㠃䛜㏣ຍ䛥䜜䜎䛩䚹

Amplification Plot ⏬㠃䠖Baseline 䛸 Threshold Line 䛾☜ㄆタᐃ䜢⾜䛔䜎䛩䚹Standard Curve ⏬㠃䠖᳨㔞⥺䛾☜ㄆ䜢⾜䛔䜎䛩䚹Gene Expression ⏬㠃䠖Target ẖཬ䜃 Sample ẖ䛾┦ᑐ್䛜⟬ฟ䛥䜜䚸䜾䝷䝣⾲♧䛥䜜䜎䛩䚹
1.

Plot Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䚸”RQ vs Sample”䠄䝃䞁䝥䝹ẖ䛾⾲♧䠅ཬ䜃”RQ vs Target”䠄䝍䞊䝀䝑䝖ẖ䛾⾲♧䠅䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

2.

Graph Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶⦪㍈⾲♧䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

3.

Orientation 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䜾䝷䝣䛾ྥ䛝䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

4.

View Replicate Results Table 䛻ྛ䝃䞁䝥䝹䛾᝟ሗ䛜グ㍕䛥䜜䛶䛔䜎䛩䚹䈄

”Normalized Qty Mean”䛜ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛷⿵ṇ䛧䛯䝍䞊䝀䝑䝖䛾್䛷䛩䚹䈄

”RQ”䛜 Plate Setup 䛷タᐃ䛧䛯 Reference Sample 䛸䛾┦ᑐ್䛷䛩䚹Reference Sample 䛾 RQ ್䛿ᚲ䛪”1” 䛻䛺䜚䜎䛩䚹
1

2

3

4

䈄

- 47 -

JP-886̲本文̲0707se.indd 46

15/07/08 17:33

Comparative CT (ǻǻCT) ᪂つᐇ㦂⣔䛾సᡂ : Advanced Setup 䜢⏝䛔䛯᪉ἲ
Advanced Setup 䛛䜙 Comparative CT (ǻǻCT)䜢㑅ᢥ䛧䛯᫬䛾タᐃ᪉ἲ䛻䛴䛔䛶ㄝ᫂䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Standard Curve 䛾タᐃ䛸␗䛺䜛㡯┠䛻䛴䛔䛶䛤ㄝ᫂䛧䜎䛩䚹䛭䛾௚䛾㡯┠䛾ゎᯒ᪉ἲ䛻䛴䛝䜎䛧䛶䛿䚸Standard Curve䛷䛾ヱᙜ㡯┠䜢䛤ཧ↷ୗ䛥䛔䚹

Experiment Properties ⏬㠃䠖1.

ྛ㡯┠䛷౑⏝ᶵჾ䚸౑⏝ヨ⸆䚸཯ᛂ䝇䝢䞊䝗➼䜢ṇ䛧䛟タᐃ䛧䜎䛩䚹䛣䛾᫬䚸”What type of Experiment do you
want to set up?”䛷䛿䚸Quantitation - Comparative CT (ǻǻCT) 䜢㑅ᢥ䛧䜎䛩䚹- 48 -

JP-886̲本文̲0707se.indd 47

15/07/08 17:33

Plate Setup ⏬㠃䠖
Define Target and Samples 䝍䝤䛷1.

䠰䠽䡎䡃䡁䡐䛸 Sample 䛾タᐃ䜢⾜䛔䜎䛩䚹

Option: 㞟ᅋ㛫䛷䛾ẚ㍑ᐃ㔞䜢⾜䛔䛯䛔ሙྜ䛿䚸㞟ᅋ䛤䛸䛾ྡ๓(Biological Replicate*) 䛾ධຊ䜒⾜䛔䜎䛩䚹Biological
Replicate 䛸䛿䚸ྠ୍䛾⏕≀Ꮫⓗ䛺㞟ᅋ䛾ᣦᐃ䛻┦ᙜ䛧䜎䛩䚹¾

Define Biological Replicate Groups 䛻 Biological Replicate 䜢Ⓩ㘓䛧䜎䛩䚹ὀグ䠖Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢 Study 䛷ゎᯒ䛧䛯䛸䛝䛻䛾䜏䚸ゎᯒ⤖ᯝ䛻཯ᫎ䛥䜜䜎䛩䚹Standard Curveཬ䜃 Relative Standard Curve 䝗䜻䝳䝯䞁䝖䛷䛿䚸Biological Replicate ゎᯒ䛿䛷䛝䜎䛫䜣䚹a. Add Biological Group 䜢䜽䝸䝑䜽䛩䜛䛸䝸䝇䝖䛻᪂つ䛾 Biological Group 䛜㏣ຍ䛥䜜䜎䛩䚹b. Biological Group Name 䜢ධຊ䛧䚸Color 䜢㑅ᢥ䛧䜎䛩䚹ab㻌 㻌 㻌 㻌 㻌 Biological Replicate 䛸䛿䚸ྠ୍䛾⏕≀Ꮫⓗ䛺㞟ᅋ䛾ᣦᐃ䛻┦ᙜ䛧䜎䛩䚹ᚑ᮶䛾䝃䞁䝥䝹㛫䛷䛾ẚ㍑䛷↓䛟䚸ᑐ㇟䜾䝹䞊䝥㛫䛷䛾ẚ㍑ᐃ㔞ゎᯒ䜢⾜䛖䛣䛸䛜ྍ⬟䛸䛺䜚䜎䛩䚹ィ⟬䛾ᴫせ䛻䛴䛝䜎䛧䛶䛿ୗ⾲䜢䛤ཧ↷ୗ䛥䛔䚹- 49 -

JP-886̲本文̲0707se.indd 48

15/07/08 17:33

Assign Targets and Samples 䝍䝤䛷ྛ䜴䜵䝹䛻 Target ᝟ሗ䛸 Sample ᝟ሗ䜢タᐃ䛧䜎䛩䚹
1.

Assign target to the selected wells : Unknown ཪ䛿 NTC 䜢ᣦᐃ䛧䜎䛩䚹

2.

Assign sample to the selected wells: Target 䛾✀㢮䛻㛵ಀ䛺䛟ྠ䝃䞁䝥䝹䛻䛿ྠ୍ྡ䜢㑅ᢥ䛧䜎䛩䚹

3.

Assign sample of selected well to biological group:㻌 䠄option: Biological Replicate ゎᯒ䜢⾜䛔䛯䛔ሙྜ䛾䜏タᐃ䠅 ྠ୍䜾䝹䞊䝥䛻ᒓ䛩䜛䝃䞁䝥䝹䜴䜵䝹䜢㑅ᢥ䛧䛶䚸ヱᙜ䛩䜛䜾䝹䞊䝥ྡ䛾 Assign 䜢䝏䜵䝑䜽䛧䜎䛩䚹

4.

Select relative quantitation settings:a.

Reference Sample 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛷┦ᑐᐃ㔞ゎᯒ䛾ᇶ‽䛸䛺䜛䝃䞁䝥䝹䜢㑅ᢥ䛧䜎䛩䚹b.

Endogenous Control 䝥 䝹 䝎 䜴 䞁 䝯 䝙 䝳 䞊 䛷 ෆ ᅾ ᛶ 䝁 䞁 䝖 䝻 䞊 䝹 䛸 䛩 䜛 㑇 ఏ Ꮚ ྡ 䜢 㑅 ᢥ 䛧 䜎 䛩 䚹 㻌 㻌 㻌 㻌 㻌 㻌 㻌 㻌 㻌 㻌 㻌䛣䛾㝿䚸Multiple Controls 䜢㑅ᢥ䛩䜛䛸」ᩘ䛾㑇ఏᏊ䜢ෆᅾᛶ䝁䞁䝖䝻䞊䝹㑇ఏᏊ䛸䛧䛶ゎᯒ䛩䜛䛣䛸䜒ྍ⬟䛷䛩䚹㑅ᢥ䛧䛯㑇ఏᏊ⤖ᯝ䛾ᖹᆒ್䛜⿵ṇィ⟬䛻౑䜟䜜䜎䛩䚹౑⏝䛧䛯䛔㑇ఏᏊ䜢㑅ᢥ䛧䠫䠧䜢䜽䝸䝑䜽䛧䜎䛩䚹

1

2

3

4a
4b

- 50 -

JP-886̲本文̲0707se.indd 49

15/07/08 17:33

Comparative CT (ǻǻCT) 䝕䞊䝍䛾ゎᯒ : ༢⊂䝗䜻䝳䝯䞁䝖ෆ䛷䛾ゎᯒὀグ䠖䛣䛣䛷䛿 Standard Curve 䛾タᐃ䛸␗䛺䜛㡯┠䛾䜏䛤ㄝ᫂⮴䛧䜎䛩䚹䛭䛾௚䛾㡯┠䛾ゎᯒ᪉ἲ䛻䛴䛝䜎䛧䛶䛿䚸Standard
Curve 䛷䛾ヱᙜ㡯┠䜢䛤ཧ↷ୗ䛥䛔䚹Comparative CT (ǻǻCT)䜢㑅ᢥ䛧䛯ሙྜ䚸Analysis ⏬㠃䛻 Gene Expression ⏬㠃䛜㏣ຍ䛥䜜䜎䛩䚹Gene Expression⏬㠃䛷䛿䚸Target ẖཬ䜃 Sample ẖ䛾┦ᑐ್䛜⟬ฟ䛥䜜䚸䜾䝷䝣⾲♧䛥䜜䜎䛩䚹

Amplification 䠬䡈䡋䡐 ⏬㠃䠖Baseline 䛸 Threshold Line 䛾☜ㄆ䜢⾜䛔䜎䛩䚹Gene Expression ⏬㠃䠖
1.

Plot Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䚸”RQ vs Sample”䠄䝃䞁䝥䝹ẖ䛾⾲♧䠅ཬ䜃”RQ vs Target”䠄䝍䞊䝀䝑䝖ẖ䛾⾲♧䠅䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

2.

Graph Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶⦪㍈⾲♧䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

3.

Orientation 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䜾䝷䝣䛾ྥ䛝䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

4.

View Technical Replicates 䛻ྛ䝃䞁䝥䝹䛾᝟ሗ䛜グ㍕䛥䜜䛶䛔䜎䛩䚹xྛ䠰䠽䡎䡃䡁䡐䛛䜙 Endogenous Control 䛾 CT ್䜢ᘬ䛔䛯್䛜 ǻCT 䛷䛩䚹xྛ䝃䞁䝥䝹䛾 ǻCT 䛛䜙 Reference Sample 䛾 ǻCT ್䜢ᘬ䛔䛯್䛜 ǻ ǻCT 䛷䛩䚹

x”RQ”䛜 Reference Sample 䛸䛾┦ᑐ್䛷䛩䚹Reference Sample 䛾 RQ ್䛿䛛䛺䜙䛪”1”䛻䛺䜚䜎䛩䚹
1

2

3

4

- 51 -

JP-886̲本文̲0707se.indd 50

15/07/08 17:33

Comparative CT (ǻǻCT) 䝕䞊䝍䛾ゎᯒ : Study 䜢⏝䛔䛯༢⊂ཪ䛿」ᩘᯛ䛾ゎᯒ」ᩘ䛾 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢 Study 䛻䛶ྠ᫬ゎᯒ䛩䜛᪉ἲ䛻䛴䛔䛶ㄝ᫂䛧䜎䛩䚹ὀグ䠖䛣䛣䛷䛿 Standard Curve 䛾タᐃ䛸␗䛺䜛㡯┠䛾䜏䛤ㄝ᫂⮴䛧䜎䛩䚹䛭䛾௚䛾㡯┠䛾ゎᯒ᪉ἲ䛻䛴䛝䜎䛧䛶䛿䚸Standard
Curve 䛷䛾ヱᙜ㡯┠䜢䛤ཧ↷ୗ䛥䛔䚹

㔜せ䠖」ᩘ䛾 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢 Study 䛻䛶ྠ᫬ゎᯒ䛩䜛ሙྜ䚹඲䛶䛾 Comparative CT (ǻǻCT)䝗䜻
䝳䝯䞁䝖䛾䝃䞊䝬䝹䝃䜲䜽䝸䞁䜾᮲௳䠄䝷䞁䝰䞊䝗䚸䝇䝔䝑䝥䚸䝃䜲䜽䝹ᩘ䚸཯ᛂᐜ㔞䠅䛜ྠ୍䛷䛒䜛䛣䛸䛜ᚲ㡲䛷䛩䚹1 䛴䛷䜒␗䛺䜛タᐃ䛜䛒䜛䛸䚸ྠ᫬䛻ゎᯒ䛩䜛䛣䛸䛜䛷䛝䜎䛫䜣䛾䛷䛤ὀពୗ䛥䛔䚹
㔜せ䠖」ᩘ䛾 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢 Study 䛻䛶ྠ᫬ゎᯒ䛩䜛ሙྜ䛻䛚䛔䛶䜒䚸ྛ䝃䞁䝥䝹䛾ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛿ྠ୍䝗䜻䝳䝯䞁䝖ୖ䛻ྵ䜎䜜䛶䛔䜛䛣䛸䛜ᚲせ䛷䛩䚹

1. 䝩䞊䝮⏬㠃䜘䜚 Create Study 䜢䜽䝸䝑䜽䛧᪂つ䝗䜻䝳䝯䞁䝖䜢㛤䛝䜎䛩䚹- 52 -

JP-886̲本文̲0707se.indd 51

15/07/08 17:33

Study Properties ⏬㠃䠖Create Study 䜢㑅ᢥ䛩䜛䛸䚸ึ䜑䛻 Study Properties ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹䛣䛾⏬㠃䛷䛿䚸Study 䝗䜻䝳䝯䞁䝖ྡ䛾ධຊ䛸䚸ゎᯒ䜢⾜䛖 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䛾㏣ຍ䜢⾜䛔䜎䛩䚹1.

“Study Properties”ෆ䛾 Study Name 䛻ྡ๓䜢ධຊ䛧䜎䛩䠄ᚲ㡲䠅䚹䛭䛾௚䛾㡯┠䛿䜸䝥䝅䝵䞁䛷䛩䚹ὀグ䠖 ౑⏝䛷䛝䜛ᩥᏐ䛿䚸ⱥᩘᏐ䚸䝇䝨䞊䝇䚸䝝䜲䝣䞁䛚䜘䜃䜰䞁䝎䞊䝞䞊䠄䛩䜉䛶༙ゅ䠅䛷䛩䚹䛣䛾䜋䛛䛾ᩥᏐ䜢౑⏝䛩䜛䛸䝣䜯䜲䝹䛜◚ᦆ䛩䜛䛣䛸䛜䛒䜚䜎䛩䚹

2.

“Set Up Experiments”䛾 Add Experiment(s)䜢䜽䝸䝑䜽䛩䜛䛸䠫䡌䡁䡊⏬㠃䛜㛤䛝䜎䛩䚹

3.

Open ⏬㠃䛷䚸Ⓩ㘓䛩䜛඲䛶䛾 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢㑅ᢥ䛧䚸Open 䜢䜽䝸䝑䜽䛩䜛䛸䚸ୗẁ䛾䝸䝇䝖䛻 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䛜⾲♧䛥䜜䜎䛩䚹

4.

䝸䝇䝖䛾䝗䜻䝳䝯䞁䝖䜢䜽䝸䝑䜽䛧䛶㑅ᢥ䛩䜛䛸䚸Properties 䛾㡯┠䛻䛭䛾䝥䝺䞊䝖䛻Ⓩ㘓䛥䜜䛶䛔䜛౑⏝䜴䜵䝹䚸Sample䚸ཬ䜃 Target 䛾᝟ሗ䜢☜ㄆ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

1

2

4
3

- 53 -

JP-886̲本文̲0707se.indd 52

15/07/08 17:33

Define Replicates ⏬㠃 䠖
Define Replicates ⏬㠃䛷䛿䚸Biological Replicate Group 䛾☜ㄆ䞉タᐃ䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䛷 Biological Replicate 䜢タᐃ䛧䛯ሙྜ䛿䚸䛣䛾⏬㠃䛻⮬ືⓗ䛻⾲♧䛥䜜䜛䛾䛷䚸㐺ᐅ☜ㄆཬ䜃タᐃኚ᭦䜢⾜䛖䛣䛸䛜䛷䛝䜎䛩䚹Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䛷 Biological Replicate 䜢タᐃ䛧䛺䛛䛳䛯ሙྜ䛿䚸䛣䛾⏬㠃䛷タィ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹1.

Add Biological Group 䜢䜽䝸䝑䜽䛧䛶⾲♧䛥䜜䜛⏬㠃䛷䚸Biological Group 䜢సᡂ䞉タᐃ䛩䜛䛣䛸䛜
䛷䛝䜎䛩䚹

2.

Edit Biological Group 䜢䜽䝸䝑䜽䛧䛶⾲♧䛥䜜䜛⏬㠃䛷䚸Biological Group 䛾ෆᐜ䜢ኚ᭦䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹ὀグ䠖1 ཬ䜃 2 䛾సᴗ䛾ヲ⣽䛿[Relative Standard Curve and Comparative CT Experiments Getting
Started Guide] 䠄ⱥᩥ䠅䜢䛤ཧ↷ୗ䛥䛔䚹

3.

䝸䝇䝖䜢䜽䝸䝑䜽䛧䛶㑅ᢥ䛩䜛䛸䚸Properties 䛻Ⓩ㘓䛥䜜䛶䛔䜛 Sample ཬ䜃 Target ᝟ሗ䛜⾲♧䛥䜜䜎
䛩䚹
1

2

3

- 54 -

JP-886̲本文̲0707se.indd 53

15/07/08 17:33

Analysis Settings⏬㠃ྑୖ䚸Analysis Settings 䜢䜽䝸䝑䜽ཪ䛿 Analysis 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛛䜙 Analysis Settings…䜢㑅ᢥ䛧䚸
Relative Quantitation Settings 䝍䝤䜢䜽䝸䝑䜽䛧䚸ゎᯒ᮲௳䜢タᐃ䛧䜎䛩䚹
1.

Reference Sample(s) 䛻䛶䚸┦ᑐ್䜢⟬ฟ䛩䜛䛯䜑䛾ᇶ‽䛸䛺䜛䝃䞁䝥䝹䜢㑅ᢥ䛧䜎䛩䚹䝋䝣䝖䜴䜵䜰䛿䛣䛣䛷タᐃ䛥䜜䛯䝃䞁䝥䝹䛾್䜢”1”䛸䛧䛯䛸䛝䛾┦ᑐ್䜢⟬ฟ䛧䜎䛩䚹
I.

Biological Replicate Group 䛾 Reference Sample 䜢ᣦᐃ䛩䜛䝥䝹䝎䜴䞁䝯䝙䝳䞊 䠄Biological
Replicate Group 䜢タᐃ䛧䛶䛔䛺䛔䛸䛝䛻䛿䝯䝙䝳䞊㑅ᢥ୙ྍ䠅

II.

Technical Replicate Group 䛿䚸Ⓩ㘓䛥䜜䛶䛔䜛඲䛶䛾 Sample 䛛䜙 Reference Sample 䜢ᣦᐃ䛩䜛䝥䝹䝎䜴䞁䝯䝙䝳䞊

2.

Endogenous Control(s) 䛻䛶䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛸䛧䛶౑⏝䛩䜛 Target 䜢㑅ᢥ䛧䜎䛩䚹䝋䝣䝖䜴䜵䜰䛿䛣䛣䛷タᐃ䛧䛯 Target 䛾ᐃ㔞್䜢⏝䛔䛶䚸䝃䞁䝥䝹ẖ䛻⮬ື⿵ṇ䜢⾜䛔䜎䛩䚹Endogenous Control 䛷 Multiple
Controls 䜢㑅ᢥ䛩䜛䛸䚸ෆᅾᛶ䝁䞁䝖䝻䞊䝹䛸䛧䛶౑⏝䛩䜛 Target 䛾㑅ᢥ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹䛣䛣䛻䝏䜵䝑䜽䜢ධ䜜䛯 Target ⩌䛾ᖹᆒ್䛜ෆᅾᛶ䝁䞁䝖䝻䞊䝹್䛸䛧䛶⿵ṇィ⟬䛻⏝䛔䜙䜜䜎䛩䚹3.

Efficiency 䛷䛿䚸ྛ Target 䛾 PCR ຠ⋡䛜᪤䛻᫂☜䛺䛸䛝䛻䚸Target ẖ䛾 PCR ຠ⋡䜢ධຊ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹䛣䛣䛻 PCR ຠ⋡䜢ධຊ䛧䛯᫬䛻䛿䚸Relative Standard Curve 䛾ゎᯒ䜰䝹䝂䝸䝈䝮䜢⏝䛔䛯┦ᑐ್䛾⟬ฟ䛜⾜䜟䜜䜎䛩䚹

4.

RQ Min/Max Calculations 䛻䛚䛔䛶䚸┦ᑐ್䛻ᑐ䛩䜛䜶䝷䞊䝞䞊䛾ィ⟬䜢 Confidence level 䜢⏝䛔䜛䛛䚸
Standard Deviations 䜢⏝䛔䜛䛛㑅ᢥ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹5.

タᐃ䛜⤊஢䛧䛯䜙 Analyze 䜢䜽䝸䝑䜽䛧䜎䛩䚹

II

I
1

2

3

4

5

- 55 -

JP-886̲本文̲0707se.indd 54

15/07/08 17:33

Amplification Plot ⏬㠃Study Menu 䛾 Amplification Plot ⏬㠃䛷ቑᖜ᭤⥺䜢☜ㄆ䛧䚸ゎᯒ᮲௳䛾☜ㄆཬ䜃ኚ᭦䜢⾜䛔䜎䛩䚹
1.

Experiment Data 䛻䛿Ⓩ㘓䛥䜜䛶䛔䜛 Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䛜⾲♧䛥䜜䜎䛩䚹඲䛶䛾Comparative CT (ǻǻCT)䝗䜻䝳䝯䞁䝖䜢䝝䜲䝷䜲䝖䛷㑅ᢥ䛩䜛䛸䚸඲䛶䛾ቑᖜ᭤⥺䛜 Plot View ⏬㠃䛻⾲♧䛥䜜䜎䛩䚹 Target 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢ษ䜚᭰䛘䛶䚸Threshold Line ཬ䜃 Baseline 䛾タᐃ☜ㄆཬ䜃ኚ᭦䜢⾜䛔䜎䛩䚹

2. ⏬㠃ྑୗ䛾 Well Results Data 䛷䛿 View Well Table 䛸 View Plate Layout 䜢ษ䜚᭰䛘䛶⾲♧䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹View Well Table 䛷䛿ᚲせ䛻ᛂ䛨䛶 Omit 䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

1

2- 56 -

JP-886̲本文̲0707se.indd 55

15/07/08 17:33

Gene Expression ⏬㠃Gene Expression ⏬㠃䛷 Sample ẖཬ䜃 Biological Replicate Group ẖ䛾┦ᑐ್䜢☜ㄆ䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹⟬ฟ䛥䜜䛯┦ᑐ್䛿 View Plot ⏬㠃䛻䜾䝷䝣⾲♧䛥䜜䜎䛩䚹
1.

Plot Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䚸”RQ vs Sample”䠄䝃䞁䝥䝹ẖ䛾⾲♧䠅ཬ䜃”RQ vs Target”
䠄䝍䞊䝀䝑䝖ẖ䛾⾲♧䠅䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

2.

Graph Type 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶⦪㍈⾲♧䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

3.

Orientation 䝥䝹䝎䜴䞁䝯䝙䝳䞊䜢⏝䛔䛶䜾䝷䝣䛾ྥ䛝䜢ษ䜚᭰䛘䜛䛣䛸䛜䛷䛝䜎䛩䚹

4.

Replicate Results Data ⏬㠃䛻䛿┦ᑐ್䛜⾲♧䛥䜜䜎䛩䚹”RQ”䛜 55 䝨䞊䝆 1 䛷タᐃ䛧䛯 Reference
Sample 䛸䛾 ┦ᑐ್䛷䛩䚹Reference Sample 䛾 RQ ್䛿䛛䛺䜙䛪”1”䛻䛺䜚䜎䛩䚹TechnicalReplicates䠄䝃䞁䝥䝹ẖ䠅䚸Biological Replicates 䛾䛭䜜䛮䜜䛿䝍䝤䜢ษ䜚᭰䛘䜛䛣䛸䛷☜ㄆ䛷䛝䜎䛩䚹䝍䝤䜢ษ䜚᭰䛘䜛䛸䜾䝷䝣䜒⮬ືⓗ䛻ኚ䜟䜚䜎䛩䚹

5.

Endo Controls 䛾䝥䝹䝎䜴䞁䝯䝙䝳䞊䛷ෆᅾᛶ䝁䞁䝖䝻䞊䝹䜢䚸Ref Sample 䝥䝹䝎䜴䞁䝯䝙䝳䞊䛷ᇶ‽䛸䛩䜛 Sample 䜔 Biological Replicate 䜢ኚ᭦䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹

6.

䝸䝇䝖䜢䜽䝸䝑䜽䛩䜛䛸䜽䝸䝑䜽䛧䛯 Sample 䜔 Biological Replicate 䛻Ⓩ㘓䛥䜜䛶䛔䜛ྛ䜴䜵䝹䛾᝟ሗ䛜⾲♧䛥䜜䜎䛩䚹㐺ᐅ Omit 䛩䜛䛣䛸䜒䛷䛝䜎䛩䚹
1

2

3

4

5

6

- 57 -

JP-886̲本文̲0707se.indd 56

15/07/08 17:33

Compare Settings ⏬㠃Compare Settings ⏬㠃䛷䛿䚸2 䛴䛾␗䛺䜛 Analysis Settings 䛻䜘䜛ゎᯒ䜢⾜䛔䚸ྠ᫬䛻⾲♧䛧䛶ẚ㍑䛩䜛䛣䛸䛜䛷䛝䜎䛩䚹
1.

Settings Comparison 䛾 Comparison Analysis Settings ᶓ䛾 Edit 䝪䝍䞁䜢䜽䝸䝑䜽䛧䚸Analysis
Setting 䜢ኚ᭦䛩䜛䛣䛸䛜ฟ᮶䜎䛩䚹2.

ゎᯒ䝕䞊䝍䜢 Results Comparison 䛷☜ㄆ䛧䜎䛩䚹ྛ㡯┠ᮎᑿ”1”䛸”2”䛷⾲♧䛥䜜䜎䛩䚹Settings
Comparison 䛷ᕥḍ䛜”1”ྑḍ䛜”2” 䛷䛩䚹3.

Use Comparison Analysis Settings 䜢䜽䝸䝑䜽䛩䜛䛸䚸䛭䛾ḍ䛷䛾タᐃ䛜䝕䞊䝍ゎᯒ䛻཯ᫎ䛥䜜䜎
䛩䚹

2

3

1

- 58 -

JP-886̲本文̲0707se.indd 57

15/07/08 17:33

Appendix A : StepOnePlus System 䛷䛾 VeriFlex ᶵ⬟䛾タᐃ᪉ἲ
StepOnePlus 䝅䝇䝔䝮䛿 6 ಶᗘ䝉䞁䝃䞊䛷 6 ಶ䛾䝤䝻䝑䜽䜢ไᚚ䛧䛶䛚䜚䚸䛭䜜䜖䛘 PCR 䛾ྛ䝇䝔䝑䝥䛷 6 ✀㢮䛾ᗘタᐃ䜢䛩䜛䛣䛸䛜ྍ⬟䛷䛩䚹

1.

Experiment 䛾 Setup ⏬㠃䛾 Plate Setup 䛛䜙䚸Assign Targets and Samples 䝍䝤䛾 View Plate Layout 䜢⾲♧䛧䜎䛩䚹䜴䜱䝄䞊䝗䜢฼⏝䛧䛶䛔䜛ሙྜ䛿 15 䝨䞊䝆䛾䝇䝔䝑䝥䠍䠍䛾⏬㠃䛷[Edit Plate Layout]䜢䜽䝸䝑䜽
䛧䜎䛩䚹

2.

96 䜴䜵䝹⏬㠃䛾ୖ䛻䛒䜛䚸Enable VeriFlex Block 䛾䝏䜵䝑䜽䝪䝑䜽䝇䛻䝏䜵䝑䜽䜢䛔䜜䜎䛩䚹

3.

[Are you sure you want to change the VeriFlex block setting?] 䛸 WARNING 䛜⾲♧䛥䜜䜎䛩䛾䛷
YES 䜢䜽䝸䝑䜽䛧䜎䛩䚹㻌 ⦪ 6 䝤䝻䝑䜽䛻ศ๭䛩䜛㉥䛔䝷䜲䞁䛜⾲♧䛥䜜䜎䛩䚹

4.

Setup 䛾 Run Method 䜢䜽䝸䝑䜽䛧䜎䛩䚹
2

1
4

3

- 59 -

JP-886̲本文̲0707se.indd 58

15/07/08 17:33

5.

タᐃኚ᭦䛧䛯䛔

ᗘ䛻䝫䜲䞁䝍䜢ྜ䜟䛫䜽䝸䝑䜽䛧䜎䛩䚹Set Temperatures ⏬㠃䛜⌧䜜䜎䛩䚹

6.

[Set different temperatures for one or more zones] 䛻䝏䜵䝑䜽䜢ධ䜜䚸ྛ䝌䞊䞁(Zone)䛻

ᗘ䜢ධຊ䛧䜎䛩䚹

ὀグ䠖㞄䜚ྜ䛳䛯䝌䞊䞁䛻 5䉝䜘䜚኱䛝䛔タᐃ䜢䛩䜛䛣䛸䛿䛷䛝䜎䛫䜣䚹

7.

OK 䜢䜽䝸䝑䜽䛧䛯䜙 VeriFlex

ᗘタᐃ䛾᏶஢䛷䛩䚹658.

ᗘ⾲♧䛜 Zones 䛻ኚ䜟䜚䜎䛩䚹- 60 -

JP-886̲本文̲0707se.indd 59

15/07/08 17:33

Appendix B : 䝍䝑䝏䝟䝛䝹᧯స᪉ἲPCĺStepOne 䝅䝇䝔䝮䜈䛾䝣䜯䜲䝹䛾㌿㏦䛸䝷䞁

1. StepOne 䝅䝇䝔䝮௜ᒓ PC 䛷䝣䜯䜲䝹䜢సᡂ䛧䚸ಖᏑ䛧䜎䛩䚹
2. StepOne 䝋䝣䝖䜴䜵䜰⏬㠃ୖ䛾 Send Experiment to instrument …䜢䜽䝸䝑䜽䛧䜎䛩䚹

3. Send Experiment to Instrument ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹䐟

Browse 䝪䝍䞁䜢䜽䝸䝑䜽䛧䚸䠍䠊䛷ಖᏑ䛧䛯䝣䜯䜲䝹䜢㑅ᢥ䛧䜎䛩䚹䐠

Send Experiment 䝪䝍䞁䜢䜽䝸䝑䜽䛧䛶 StepOne 䝅䝇䝔䝮䜈䝣䜯䜲䝹䜢㌿㏦䛧䜎䛩䚹

4. Main Menu ⏬㠃ෆ䛾 Browse / New Experiments 䜢ᢲ䛧䜎䛩䚹- 61 -

JP-886̲本文̲0707se.indd 60

15/07/08 17:33

5. Browse Last Accessed Experiments ⏬㠃ෆ䛻㌿㏦䛧䛯䝣䜯䜲䝹䛜⌧䜜䜎䛩䚹䠆 䝣䜯䜲䝹䜢㑅ᢥ䛧䚸Copy 䝪䝍䞁䜢ᢲ䛩䛸 Template 䛸䛧䛶ಖᏑ䛩䜛䛣䛸䜒ฟ᮶䜎䛩䚹ಖᏑඛ䝣䜷䝹䝎䜢
Template 䛻ᣦᐃ䛧䚸Save & Exit 䜢ᢲ䛧䛶䛟䛰䛥䛔䚹Template 䛸䛧䛶ಖᏑ䛧䛯䝣䜯䜲䝹䜢⏝䛔䛶䝷䞁䜢⾜䛖᪉ἲ䛿ḟ䝨䞊䝆䜢ཧ↷䛧䛶䛟䛰䛥䛔䚹

6. 䝥䝺䞊䝖䜢 StepOne㻌 䝅䝇䝔䝮䛻䝉䝑䝖ᚋ䚸䝣䜯䜲䝹䜢㑅ᢥ䛧䚸⏬㠃ᕥୗ䛾 Start Run 䜢ᢲ䛧䛶䝷䞁䜢㛤ጞ䛧䜎䛩䚹

7. 䝷䞁䛜⤊஢䛩䜛䛸⮬ືⓗ䛻 Main menu ⏬㠃䛻ᡠ䜚䜎䛩䚹USB 䝯䝰䝸䜢䝫䞊䝖䛻ᤄ䛧䚸Collect Results 䜢ᢲ䛧䜎䛩䚹䝣䜯䜲䝹䛜㌿㏦䛥䜜䜎䛩䛾䛷䚸㌿㏦᏶஢ᚋ OK 䜢ᢲ䛧䛶 USB 䝯䝰䝸䜢ᢤ䛔䛶䛟䛰䛥䛔䚹䛣䛾ᚋ䛿 PC ୖ䛷䛾ゎᯒసᴗ䛸䛺䜚䜎䛩䚹ὀ䠖䜴䜵䝹タᐃ῭䜏䛾䝣䜯䜲䝹䜢 PC 䛛䜙㌿㏦䛧䛶䝷䞁䛩䜛ሙྜ(Template 䛸䛧䛶ಖᏑ䛧䛯ሙྜ䜒ྵ䜐)䚸䜴䜵䝹タᐃ῭䜏䛾䝣䜯䜲䝹䛻䛺䛳䛶䛔䜎䛩䚹- 62 -

JP-886̲本文̲0707se.indd 61

15/07/08 17:33

ಖᏑ Template 䝣䜯䜲䝹䜢࿧䜃ฟ䛩᪉ἲ

1. 䠄ึᅇ䛾䜏䠅Main Menu ⏬㠃䛷䛔䛪䜜䛛䛾 Shortcut 䝪䝍䞁䜢ᢲ䛧䜎䛩䚹Configure Shortcuts ⏬㠃䛜⌧䜜䜛䛾䛷䚸
3 䝨䞊䝆┠䛾 Create New Experiment 䜢㑅ᢥ䛧䛶 Done 䜢ᢲ䛧䜎䛩䚹㻌 䠄ḟᅇ䛛䜙䛿䛣䛾᧯స䛿ᚲせ䛒䜚䜎䛫䜣䚹䝇䝔䝑䝥 2. 䛛䜙⾜䛳䛶䛟䛰䛥䛔䚹䠅㻌
2. Main Menu ⏬㠃䛾 Create New Experiments 䜢ᢲ䛧䜎䛩䚹

3. Create an Experiment ⏬㠃䛜㛤䛝䜎䛩䚹䛣䛾୰䛛䜙௵ព䛾䝣䜯䜲䝹䜢㑅ᢥ䛧䚸⏬㠃ᕥୗ䛾 Select 䜢ᢲ䛧䜎䛩䚹

4. PCR 䝥䝻䜾䝷䝮䛜⾲♧䛥䜜䜎䛩䚹☜ㄆᚋ䚸⏬㠃ྑୗ䛾 Save 䜢ᢲ䛧䜎䛩䚹- 63 -

JP-886̲本文̲0707se.indd 62

15/07/08 17:33

5. ಖᏑ⏬㠃䛜⾲♧䛥䜜䜎䛩䚹Reaction Volume 䜢☜ㄆ䛧䚸Save & Exit 䜢ᢲ䛧䜎䛩䚹

6. Browse Last Accessed Experiments ⏬㠃ෆ䛻ಖᏑ䛧䛯䝣䜯䜲䝹䛜⾲♧䛥䜜䜎䛩䚹㻌 䝥䝺䞊䝖䜢 StepOne 䝅䝇䝔䝮䛻䝉䝑䝖ᚋ䚸䝣䜯䜲䝹䜢㑅ᢥ䛧䚸⏬㠃ᕥୗ䛾 Start RUN 䜢ᢲ䛧䛶䝷䞁䜢㛤ጞ䛧䜎䛩䚹

7. 䝷䞁䛜⤊஢䛩䜛䛸⮬ືⓗ䛻 Main Menu ⏬㠃䛻ᡠ䜚䜎䛩䚹USB 䝯䝰䝸䜢䝫䞊䝖䛻ᤄ䛧䚸Collect Results 䜢ᢲ䛧䜎䛩䚹䝣䜯䜲䝹䛜㌿㏦䛥䜜䜎䛩䛾䛷䚸㌿㏦᏶஢ᚋ OK 䜢ᢲ䛧䛶 USB 䝯䝰䝸䜢ᢤ䛔䛶䛟䛰䛥䛔䚹䛣䛾ᚋ䛿 PC ୖ䛷䛾ゎᯒసᴗ䛸䛺䜚䜎䛩䚹ὀ䠖StepOne 䝅䝇䝔䝮䛻䝕䝣䜷䝹䝖䛷ධ䛳䛶䛔䜛䝥䝻䜾䝷䝮䛷䝷䞁䜢⾜䛳䛯ሙྜ䚸ゎᯒ᫬䛻䝋䝣䝖䜴䜵䜰ୖ䛷䜴䜵䝹タᐃ䛸⾜䛖ᚲせ䛜䛒䜚䜎䛩䚹
ὀ䠖PC 䛛䜙㌿㏦䛧䛯䝣䜯䜲䝹䜢 Template 䛸䛧䛶ಖᏑ䛧䚸䛭䛾䝣䜯䜲䝹䜢࿧䜃ฟ䛧䛶䝷䞁䜢⾜䛳䛯ሙྜ䠄61 䝨䞊䝆䠅䚸䜴䜵䝹タᐃ῭䜏䛾䝣䜯䜲䝹䛻䛺䛳䛶䛔䜎䛩䚹ゎᯒ᫬䛻䜴䜵䝹タᐃ䜢⾜䛖ᚲせ䛿䛒䜚䜎䛫䜣䚹- 64 -

JP-886̲本文̲0707se.indd 63

15/07/08 17:33

Appendix C : ᮏయ䛛䜙䝕䞊䝍䜢ᅇ཰䛩䜛᪉ἲ㻌
StepOne 䛚䜘䜃 StepOnePlus ᮏయ䛿䚸᭱ᚋ䛻⾜䛳䛯䝷䞁䝕䞊䝍䜢ಖᣢ䛧䛶䛔䜎䛩䚹ୗグ䛾䜘䛖䛺䜿䞊䝇䛷䚸ᮏయ䛛䜙䝕䞊䝍䜢ᅇ཰䛩䜛䛣䛸䜢䛚່䜑䛧䜎䛩䚹䞉䝷䞁୰䛻 PC 䛜䝅䝱䝑䝖䝎䜴䞁䛧䛯䜚䚸䝋䝣䝖䜴䜵䜰䛜䝣䝸䞊䝈䛩䜛䛺䛹䛾䝖䝷䝤䝹䛜Ⓨ⏕䛧䚸PC 䛸䛾䝁䝭䝳䝙䜿䞊䝅䝵䞁䛜ษ䜜䛶䛧䜎䛳䛯ሙྜ䞉䝷䞁⤊஢ᚋ䚸䝣䜯䜲䝹䛜㛤䛛䛺䛔➼䛾ၥ㢟䛜Ⓨ⏕䛧䛯ሙྜ䞉䝷䞁㛤ጞ┤ᚋ䛻 Stop 䝪䝍䞁䜢䜽䝸䝑䜽䛧䚸䝷䞁䜢Ṇ䜑䛯ሙྜ䚹䛣䛾ሙྜ䚸ᮏయୖ䛻䝣䜯䜲䝹䛜ṧ䛳䛶䛧䜎䛔䚸୍ᗘ䝕䞊䝍ᅇ཰సᴗ䜢⾜䜟䛺䛔䛸ḟ䛾䝷䞁䛜㛤ጞ䛷䛝䛺䛔䛯䜑ᮏయ䛛䜙䝕䞊䝍䜢ᅇ཰䛩䜛᪉ἲ䛿 2 ㏻䜚䛒䜚䜎䛩䚹

StepOne 䝋䝣䝖䜴䜵䜰⤒⏤䛷䝕䞊䝍ᅇ཰䛩䜛᪉ἲ

1.

ᮏయ䛾㟁※䜢ධ䜜䚸䝍䝑䝏䝟䝛䝹䛜ᢲ䛫䜛≧ែ䛻䛺䛳䛶䛔䜛䛣䛸䜢☜ㄆ䛧䜎䛩䚹

2.

PC 䛾㟁※䜢ධ䜜䜎䛩䚹

3.

䝋䝣䝖䜴䜵䜰䜢㉳ື䛧䜎䛩䚹

4.

䝋䝣䝖䜴䜵䜰⏬㠃ୖẁ䛾 Download Experiment From Instrument 䜢䜽䝸䝑䜽䛧䜎䛩䚹

5.

2 䛾 Browse 䜢䜽䝸䝑䜽䛧䚸䝣䜯䜲䝹䛾䝎䜴䞁䝻䞊䝗ඛ䜢㑅ᢥ䛧䚸Download Experiment 䜢䜽䝸䝑䜽䛧䜎䛩䚹

6.

䝎䜴䞁䝻䞊䝗䛜᏶஢䛩䜛䛸䚸᏶஢䛧䛯䛣䛸䜢♧䛩⏬㠃䛜⾲♧䛥䜜䜎䛩䚹ὀグ䠖ṇᖖ䛻䝷䞁䛜⤊஢䛧䛯䝣䜯䜲䝹䛾ሙྜ䚸䝎䜴䞁䝻䞊䝗䛜᏶஢䛩䜛䜎䛷ᩘศ䛛䛛䜛䛣䛸䛜䛒䜚䜎䛩䚹᏶஢䜢♧䛩⏬
㠃䛜⾲♧䛥䜜䜛䜎䛷䚸䝋䝣䝖䜴䜵䜰䜢᧯స䛫䛪䛻䛭䛾䜎䜎䛚ᚅ䛱ୗ䛥䛔䚹7.

䝎䜴䞁䝻䞊䝗䛧䛯䝣䜯䜲䝹䜢㛤䛝䚸ෆᐜ䜢☜ㄆ䛧䜎䛩䚹- 65 -

JP-886̲本文̲0707se.indd 64

15/07/08 17:33

ᮏయ䛛䜙 USB 䝯䝰䝸䛷┤᥋ᅇ཰䛩䜛᪉ἲ

1.

ᮏయ䛾㟁※䜢ධ䜜䚸䝍䝑䝏䝟䝛䝹䛜ᢲ䛫䜛≧ែ䛻䛺䛳䛶䛔䜛䛣䛸䜢☜ㄆ䛧䜎䛩䚹

2.

USB 䝯䝰䝸䜢䝫䞊䝖䛻䛥䛧䜎䛩䚹⏬㠃ୗẁ䛻 USB 䛾䝬䞊䜽䛜⾲♧䛥䜜䜛䛾䜢☜ㄆ䛧䜎䛩䚹

3.

⏬㠃ᕥୖ䛾 Collect Results 䜢ᢲ䛧䜎䛩䚹䝣䜯䜲䝹䛜㌿㏦䛥䜜䜎䛩䚹4.

㌿㏦᏶஢䛾䝯䝑䝉䞊䝆⏬㠃䛜⾲♧䛥䜜䛯䜙䚸OK 䜢ᢲ䛧䛶 USB 䝯䝰䝸䜢ᢤ䛝䜎䛩䚹

5.

StepOne 䝋䝣䝖䜴䜵䜰䛜䜲䞁䝇䝖䞊䝹䛥䜜䛶䛔䜛 PC ୖ䛷䝣䜯䜲䝹䜢㛤䛝䚸ෆᐜ䜢☜ㄆ䛧䜎䛩䚹- 66 -

■アプリケーションサポートについて
テクニカルサポート

TEL：0120-477-392（受付時間：月曜日〜金曜日 9:00 〜 18:00 祝日除く）

jptech@lifetech.com■修理・メンテナンスなどのアフターサービスについて
修理、点検整備、キャリブレーションサービス、バリデーション、移設などについてはこちらまでお問い合わせください。

TEL：0120-203-885 FAX：03-6832-9587（受付時間：月曜日〜金曜日 9:00 〜 18:00 祝日除く）■保守契約について
TEL：0120-203-885 FAX：03-6832-9588（受付時間：月曜日〜金曜日 9:00 〜 12:00、13:00 〜 17:30 祝日除く）

www.thermofisher.com/steponeplus

研究用にのみ使用できます。診断目的およびその手続上での使用はできません。
記載の社名および製品名は、弊社または各社の商標または登録商標です。標準販売条件はこちらをご覧ください。www.lifetechnologies.com/TC

For Research Use only.Not for use in diagnostic procedures. © 2015 Thermo Fisher Scientific Inc. All rights reserved.
All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. TaqMan is a registered trademark of Roche Molecular Systems, Inc.
Microsoft and Windows are registered trademarks of Microsoft Corporation. Printed in Japan, 07/2015, Part Number JP886 Rev.H

サーモフィッシャーサイエンティフィック
ライフテクノロジーズジャパン株式会社
本社：〒108-0023

東京都港区芝浦 4-2-8

jptech@lifetech.com
0120-477-392
テクニカルサポート
TEL：03-6832-6980 FAX：03-6832-9584
オーダーサポート
TEL：03-6832-9300 FAX：03-6832-9580
営 業 部