A comparison of direct (flow cytometry) and indirect (stomatal guard cell lengths and chloroplast numbers) techniques as a measure of ploidy in black wattle, Acacia mearnsii (de Wild)

Three assays, namely stomatal length measurements, counts of chloroplast numbers within the stomatal guard cells and flow cytometry, were used to confirm the ploidy of three diploid and three tetraploid Acacia mearnsii lines. The first two methods have previously been successful in identifying ploidy in this species, but they are indirect assessments. Flow cytometry directly quantifies the amount of DNA present in a sample, but has not been previously tested on A. mearnsii. It was decided to test the accuracy of all three methods by using the exact same plant material for each method. Results showed that each method correctly identified the ploidy of the diploid plant material tested as well as two of the three tetraploids tested. All three techniques identified the third tetraploid (C19/48/17) as a diploid, suggesting that its original classification was incorrect. From this study, all three techniques followed the same trend, confirming their accuracy for future research into ploidy identification of A. mearnsii. Flow cytometry has the advantage of providing a quick and efficient direct assessment of DNA in the sample, but is the most expensive. The other two techniques used (stomatal length measurements and counts of chloroplast numbers within the stomatal guard cells) are also accurate but are indirect methods and more time-consuming.