Esophageal Adenocarcinoma Methods and Protocols

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by the quality filters for being polyclonal, having low read

quality, or the result of primer dimer.

4. These reports are used to evaluate the quality of the Ion
   Torrent PGM run. A good quality run has at least 30% ISP
   loading and 30% usable reads.


5. The data from the sequencing runs can be analyzed using the
   Torrent Suite analysis pipeline, which includes raw sequencing
   data processing (DAT processing), splitting of the reads
   according to the barcode for the individual sample output
   sequence, classification, signal processing, base calling, read fil-
   tering, adapter trimming, and alignment quality control.

4 Notes

1. If you are using a DNA panel with two primer pools, set up
   two 10-μL amplification reactions, then combine them after
   target amplification to give a volume of 20 μL.


2. Cycle number recommendations in the table are based on
   10-ng DNA input. Cycle number can be increased when input
   material quality or quantity is questionable.


3. Residual ethanol drops inhibit library amplification. If needed,
   centrifuge the plate and remove remaining ethanol before air-

Fig. 5 Chip Minifuge for centrifugation of chip. It enables effective and efficient sample loading. The minifuge
quickly accelerates to maximum speed and stops quickly

Next Generation Sequencing and Esophageal Adenocarcinoma