The effect of replicate number and image analysis method on sweetpotato [Ipomoea batatas (L.) Lam.] cDNA microarray results

Microarray analysis makes it possible to determine the relative expression of thousands of genes simultaneously. It has gained popularity at a rapid rate, but many caveats remain. In an effort to establish reliable microarray protocols for sweetpotato [Ipomoea batatas (L.) Lam.], we compared the effect of replication number and image analysis software with results obtained by quantitative real-time PCR (Q-RT-PCR). Sweetpotato storage root development is the most economically important process in sweetpotato. In order to identify genes that may play a role in this process, RNA for microarray analysis was extracted from sweetpotato fibrous and storage roots. Four data sets, Spot4, Spot6, Finder4 and Finder6, were created using 4 or 6 replications, and the image analysis software of UCSF Spot or TIGR Spotfinder were used for spot detection and quantification. The ability of these methods to identify significant differential expression between treatments was investigated. The data sets with 6 replications were better at identifying genes with significant differential expression than the ones of 4 replications. Furthermore when using 6 replicates, UCSF Spot was superior to TIGR Spotfinder in identifying genes differentially expressed (18 out of 19) based on Q-RT-PCR. Our study shows the importance of proper replication number and image analysis for microarray studies.