Genotyping Example, Step 2 - Measuring The Observations
The second step is where the researchers do the lab procedures for visualizing DNA marker patterns in their testing population to compile their "observed data".
Figure 7. Sample gel photo of CAPS marker CosOH57, genotyped in an F2 population. Photo credit: Matthew Robbins, The Ohio State University
The gel photo to the right (Fig. 7) shows the results of screening F2 individuals from our same OH88119 x 6.8068 cross with the CosOH57 DNA marker. Lane 1 is the ladder, lanes 2 and 3 are DNA patterns from the parents and lanes 4 - 33 are DNA samples from the F2 individuals. Notice the 3 different patterns seen in the F2s with: 1) a pattern like parent OH88119, 2) parent 6.6068 or 3) a heterozygous pattern which combines both parental patterns (same as seen in the F1s).
Each F2 individual plant is then scored with an A, B, or H code, as outlined in Table 4 below. For more information or practice on scoring a gel, you can go to the eXtension lesson, Genotyping With Molecular Markers: Scoring a Molecular Marker on an Agarose Gel or to an interactive animation in the Plant and Soil Sciences eLibrary where you can practice scoring a gel and coding it (Gel Scoring).
| Code | Genotype |
|---|---|
| A | homozygous for parent 1 allele (OH88119 in this case) |
| B | homozygous for parent 2 allele (6.8068 in this case) |
| H | heterozygous, containing one allele from each parent |
We then total up our scoring data and enter it into a chi-square table, much like we did in the phenotyping example. The observed column is the data collected from the gel (Table 5).
| Class | Observed (O) | Expected Frequency | Expected (E) |
|---|---|---|---|
| A (OH88119) | 85 | 0.25 | 49.25 |
| B (6.8068) | 46 | 0.25 | 49.25 |
| H (heterozygous) | 66 | 0.50 | 98.50 |
| TOTAL | 197 | 1 | 197 |