Identification of triploid Citrus by isozyme analysis

Seedlessness is a desirable horticultural attribute in Citrus and is positively associated with triploidy. The conventional cytological method for triploid identification is a laborious technique involving the preparation of root tips for chromosomal analysis. Digital densitometry of isozymes, however, offers the possibility of distinguishing triploid Citrus from large populations of seedlings both quickly and cheaply. Where there are no gene dosage regulation effects, greater band density should be evident in the allozyme contributed by the diploid gamete for a heterozygous locus. The isozymes of 4 enzymes; malate dehydrogenase, 6- phosphogluconate dehydrogenase, shikimate dehydrogenase, and phosphoglucose isomerase, were investigated with polyacrylamide gel electrophoresis. Band densities of these isozymes for triploid Citrus, their diploid siblings and diploid progenitors were measured using a digital densitometer. Of the 4 enzymes investigated only allozymes for sbikimate dehydrogenase exhibited consistent differences over a wide range of Citrus cultivars. Greater band density was evident in the allozyme contributed by the diploid gamete. The band density ratio between allozymes for triploid Citrus was close to 0.5, while for diploid Citrus band density ratios were close to 1.0. This effect is due to the extra protein coded by the additional gene dose and was not observed in diploids. Shikimate dehydrogenase proved to be an accurate molecular marker for distinguishing between diploid and triploid Citrus for heterozygous progeny.