The nano-drop (Table 1) and agarose gel imaging (Figure 1) were used to determine the concentration and purity of the isolate RNA.

Table 1. Table of yeast RNA concentrations and purities.

Figure1. Gel image of purified RNA. Lane1: ZMSD1; Lane 2: WT; Lane 3: ZMSD1; Lane 4: blank;

Lane 5: ZMSD1; Lane 6: WT; Lane 7: ZMSD2; Lane 8: ZMSD2; Lane 9: blank; Lane 10: WT;

Lane 11: ZMSD2; Lanes numbers are consistent with sample numbers in Table 1.

According the concentrations and purities of the nano-drop and gel image, samples 6 and 11 were used for WT and ZMSD2 microarray hybridization (Figure 2).  The WT was dyed red and ZMSD2 was dyed green.

   A                                                                                                 B

Figure 2. Picture of microarray slides using sample 6 and 11. (A) Depicts microarray image under normal view and a gain of 10. (B) Depicts the same microarray image under ratio view and a gain of 10.
 

This microarray data was not able to be analyzed.  Therefore data from a previous class was used for analysis (Figure 3). Yeast WT genome was compared to ZMSD2 genome.  The WT samples were dyed green while ZMSD2 was dyed red.

   A                                                                                                   B

Figure 3. Picture of microarray slide from 2008 class data.  (A) Depicts microarray image under normal view and a gain of 10. (B) Depicts the same microarray image under a ratio view and a gain of 10.

The scanalyze software was used to analyze the raw data (Figure 3). The program measured the pixels of light emitted by each spot on the microarray image. This data was then imported into excel and manipulated. This involved subtracting out the background intensities and creating ratios of mutant vs. wild type.

This data was then combined with values for ZMSD1 vs. WT for comparison with ZMSD2 and WT. The dye reversal microarrays for these two sets of arrays were unable to be analyzed due to insufficient microarray slide data. This combined data was then imported into magic tool for a log base 2 transformation and subsequent mean normalization and standardization. This data was then selectively filtered based on extreme values.

Figure 4. Box and Whisker Plot: This represents the Data from ZMSD1 and ZMSD2 after log base 2 transformation and subsequent normalization and standardization. The first two plots are for ZMSD2 vs WT and the second two are from ZMSD1vsWT

Figure 5. This graph represents the genes picked for further study in the yeast genome due to their abnormal expression patterns in ZMSD1 and ZMSD2 knockouts. The genes shown above are YKR010C, YLR243W, YJR009C, YBR118W, YHR174W, YPR080W, YJL052W, and YCL019W.

Table 2. Table of the function and regulation of genes of interest based on Figure 5. A negative sign represents down regulation. Fold variation is between ZMSD2 and ZMSD1 knockouts.

Table 3. Table of the function and regulation supplemental genes of interest. A negative sign represents down regulation.

Title Page

Introduction

Methods

Discussion

References