プロトコル（PDFファイル）
Contents
1. PCR LB 1 5 mL 37 C GenElute PlasmidMiniprep Kit SIGMA PLN 70 5L uncut pTD1 PCR TOYOBO msertCheck Ready Blue PIK 251 pTD1 161 179 5 GCAGATTGTACTGAGAGTG 3 pTD1 845 827 5 GGAAACAGCTATGACCATG 3 InsertCheck Ready Blue 1 mL 150 pmol 100 M 1 5L PCR POR 10uxL PCR2. 3 85L C DNA 85uL 10 x Buffer 10uL 30 50U 100gxL 37 C 2 2 pTD1 5 g pTD1 coRV pTD1 5 g DNA 10L 10 x Buffer 5uL FcoRV 30 U 50L 37 C 1 1uL 3 kbp EcoRV Buffer
3. LB PCR PCR PCR 94C 4 min 94C 30 sec 50C 5 sec 30 cycles 72 C x sec kbp 0 7 kbp 2 kbp 30 sec 2 kbp 1 min 2 5 kbp kbp 0 7 kbp 0 7 kbp 6 LB mRNA 1 5 mL 5 g mRNA
4. 2 2007 10 Copyright 2007 Shimadzu Corp All Rights Reserved SHIMADZ BIOT EC H i SE Transdirect insect cel FcoRV pTD1 49L 10 L EdgeBioSystems 70437 100 150L 5 15000rpm 5 70 15000rpm 5 5 85L DNA 85uL 10 x Buffer 10uL 30 50U 100L 37 C 2 QIAquick PCR Purification Kit 50 QIAGEN
5. KOD plus TOYOBO KOD 201 cDNA 5ng 96C 2min 10 x Buffer 5uL 96C 15sec dNTP 2mM each 5 50C 30sec 30 25mM MgSO4 2uL 68 C 1min kbp N 10 M 1 5L 68 C 7min C 10 M 1 5L 4 C co KOD plus 1L H2O to 50L 2xL QIAquick PCR Purification Kit 50 QIAGEN 28104 48 L PCR 250L Binding Buffer BP 13000rpm 1 1 2007 10 Copyright 2007 Shimadzu Cor
6. kbp 0 7 kbp PCR mRNA TE Buffer 50L 20L 50L 5 g PCR lt gt 2007 10 Copyright 2007 Shimadzu Corp All Rights Reserved
7. 4 DH5 w E coli DH5w Competent Cells 9057 2L Competent Cells 20L 30 42 C 1 5 SOC 180L 37 C 1 100 gt LB 37 C 3 2007 10 Copyright 2007 Shimadzu Corp All Rights Reserved SHIMADZ BIOT EC H 5 Hramnsdirect insect ce 5 PCR C
8. DNA 7 PCR pTD1 20 A 2 4 2007 10 Copyright 2007 Shimadzu Corp All Rights Reserved SHIMADZl B lOTECH Transdirect insect cel pTD1 161 179 N pTD1 412 394 C 50 C O pTD1 161 179 5 GCAGATTGTACTGAGAGTG 3 pTD1 412 394 5 ACAACGCACAGAATOCTAGC 3 8 mRNA
9. 15 30 mer 2 x A T 4 x G C 55 60 PCR 5 C 5 GGGGTACCTTANNNNNNNNNN 3 N kpnl stop 2 40 mer 2x A T 4 x G C 55 60 2 3 Kpnl GG 2 Stop PCR Terminal transferase Fidelity
10. 28104 100L 500L Binding Buffer BP 13000rpm 1 750L PE 13000rpm 1 15000rpm 2 50wL Elution Buffer EB 13000rpm 1 260nm 50 L Elution buffer 70ng L 3 1 2 NEW ENGLAND Biolabs Quick Ligation Kit M2200S 10 g mL 2 L 1 10 2XQuick Ligation Reaction Buffer 10uL Quick T4 DNA Ligase 1uL 20L 25C 5
11. mRNA PCR PCR Figelity Hind o Transdirect secr ce 7 RNase 125 g mL PCR PCR Plasmid 1 ng u L 5 10 x Buffer 5uL dNTPs 2mM each 5 L 25 mM MgSO4 2uL pTD1 161 179 10 M 1 5uL pTD1 845 827 10 M 1 5uL KOD plus 1uL H O 29uL 50uL 96 C 96 C 50 C 68 C 68 C 4 C 2 min 15 sec 30 sec 30 cycles 1 min 1 kbp 7 min Co 2L
12. SHIMADZ BIOT EC H i Transdirect insect cel 1 DNA 5UTR pTD1 EcoRV Kpnt 90 EcoRV Ao EcoRV EcoRI 20 30 EcoRV gas7HI 10 Sma Xbaa 2 ON 5 ATGNNNNNNNNNNNNNNN 3 N
13. p All Rights Reserved SHIMADZ BIOT EC H i SE Transdirect insect cel 750 L PE 13000rpm 1 15000rpm 2 50L Elution Buffer EB 13000rpm 1 5 PCR 5 TOYOBO T4 Polynucleotide Kinase PNK 111 25uxL 50uxL Denaturation Buffer 35uL Denaturation Buffer 90C2 DNA 75L 35L Blunt End Kinase Buffer 10L 5uL 0 1M ATP 1L 0 5uL T4 Polynucleotide Kinase 2uL 1uL H20 12uL 8 5uL 100uL 50uL 37 C 1 90C 2
Download Pdf Manuals
Related Search