I try to sequence the RNAs from 2 conditions, 5 replicates per condition, using multiplex Illumina Hiseq 2000. I would like to consult about experiment design. There are 3 plans. Any suggestions are greatly appreciated, especially for the third one.
First, 5 samples are put in one sequencing lane (2 for condition A, 3 for condition B, and vice versa). This will be asymmetric.
Second, randomly choose 4 samples in one lane (2 for condition A, 2 for condition B). This reduce the replicates but increase the reads number for each sample.
Third, keep 5 reps each and pool all 10 and then run half a pool each on two lanes. Each lane would generate 10 separate fastq files and one could then concatenate the 2 fastq files (from the 2 lanes) for the same samples to get more reads for each sample, ending up with a total of 10 fastq files. We are not sure if there are other considerations in this scenario.
First, 5 samples are put in one sequencing lane (2 for condition A, 3 for condition B, and vice versa). This will be asymmetric.
Second, randomly choose 4 samples in one lane (2 for condition A, 2 for condition B). This reduce the replicates but increase the reads number for each sample.
Third, keep 5 reps each and pool all 10 and then run half a pool each on two lanes. Each lane would generate 10 separate fastq files and one could then concatenate the 2 fastq files (from the 2 lanes) for the same samples to get more reads for each sample, ending up with a total of 10 fastq files. We are not sure if there are other considerations in this scenario.
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