A Practical Guide to Cancer Systems Biology
16 A Practical Guide to Cancer Systems Biology Prepare FSM and SuperScript II mix at the ratio of 1μL SuperScript II for each 9 ...
Transcriptome Analysis: Library Construction 17 Incubate the sample at room temperature for 30 seconds, then remove and disca ...
18 A Practical Guide to Cancer Systems Biology Incubate the sample at room temperature for 30 seconds, then remove and discard ...
Transcriptome Analysis: Library Construction 19 Place the sample on the thermal cycler to preform adapter ligation. 30 ◦C fo ...
20 A Practical Guide to Cancer Systems Biology Note: Leave the sample tubes on the magnetic stand while performing all the follo ...
Transcriptome Analysis: Library Construction 21 of the PCR amplified library. Gently pipette the entire volume up and down 10 ...
22 A Practical Guide to Cancer Systems Biology (Optional) Transfer 1μL of the clear supernatant to a new PCR tube for library v ...
3. Quantitative Proteome: The Isobaric Tags for Relative and Absolute Quantitation (iTRAQ) Yi-Hsuan Wu and Hsueh-Fen Juan∗ Insti ...
24 A Practical Guide to Cancer Systems Biology derived from the digestion of the proteins.^4 Shotgun proteomics, also called pep ...
Quantitative Proteome 25 label-free method is quantification by comparing the intensity of mass spectrometric signal of each p ...
26 A Practical Guide to Cancer Systems Biology Combined Sample A Sample C Sample D Isobaric tag 145 Da Reporter group 114-117 Da ...
Quantitative Proteome 27 Sample A Sample B Sample C Sample D Protein extraction Large-scale experiment Small-scale experiment ...
28 A Practical Guide to Cancer Systems Biology Equipment Magnetic stir plate and stir bar Mortar and pestle 1.5 mL centrifuge t ...
Quantitative Proteome 29 200 mM S-Methyl methanethiosulfonate (MMTS) (500μL) Mix 9. 73 μL of MMTS and 490. 27 μL of Milli-Q w ...
30 A Practical Guide to Cancer Systems Biology Centrifuge the lysate at 17,000 g, 4◦C for 30 minutes. Transfer the supernatant, ...
Quantitative Proteome 31 Cut the gel into small pieces with tweezers (about 1 mm in diameter) and add 200μL of 25 mM TEAB to ea ...
32 A Practical Guide to Cancer Systems Biology Prepare the trypsin solution: (1) If you will exhaust the whole bottle containin ...
Quantitative Proteome 33 Vortex the gel pieces for 15 minutes and spin down. Transfer the supernatant to the mixture collection ...
34 A Practical Guide to Cancer Systems Biology Resuspend the dried peptides in each final peptide tube with iTRAQ Dissolution B ...
Quantitative Proteome 35 Prepare a new 2 mL protein lobind tube. Spin down the labeled peptides and combine the four samples of ...
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