waxy PCR

--David Farrar, Steve Larson and Ed Weck

The evaluation of individuals within a population is an essential aspect of any breeding program and the rapid identification of plants with desirable characterististics could save time and effort. In order to assess PCR (the Polymerase Chain Reaction) for application to plant breeding problems, a screen for important sources of the waxy gene has been developed.

The location of the waxy probe used for hybridization studies with six starchy inbreds and three waxy inbreds is shown in Figure 1. The band sizes were determined with a sonic digitizer from a Southern blot with Lambda DNA digested with HindIII as a marker. The waxy inbreds all have larger waxy fragments than the starchy inbreds.

The waxy hybridization probes were sequenced terminally and DNA primers designed therefrom. DNA amplification was compared at two temperatures, 60 C and 65 C, using the corn lines listed in Figure 1. The fragment sizes generated with PCR agree with the RFLP results (Fig. 2) except for Inbred 5 which gives a 100 bp smaller band in PCR than in hybridization. The waxy 2 inbred appears to be a null allele when amplified at 65 C suggesting that one of the DNA primer binding sites is causally related to the waxy mutation. PCR is more rapid than RFLP analysis and the resolution of small fragments on analytical gels is greater than on Southern blots.

Figure 1.,waxy Probe Location

Figure 2.,waxy PCR

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