Differentiation of nucellar and zygotic seedlings in citrus using ISSR markers

In the present study we propose the use of inter simple sequence repeat (ISSR) markers as an alternative methodology for early differentiation of zygotic and nucellar seedlings in citrus. The DNA amplification profiles of in vitro grown seedlings were compared to that of respective mother plant. Nucellar seedlings displayed a similar banding pattern as that of mother plants, whereas zygotic seedlings were differentiated from other seedlings by their different banding pattern. The primers P-7, I-9, I-10, I-13 and I-14 were able to identify the zygotic seedlings based on polymorphic bands in 'Soh sarkar'. In 'Attani', zygotic seedlings were identified using the primers P-1, P-7, P-15, I-8, I-9, I-10, I-13 and I-14. Out of 8 primers tested only 2 primers I-8 and I-14 showed polymorphism in Kinnow and could differentiate zygotic seedlings. The primers P-1, I-8, I-9, I-14 and P-15 generated polymorphism among 'Karna khatta' seedlings enabling distinction between nucellar and zygotic seedlings. In most of the cases, single ISSR primer was not able to identify all the zygotic seedlings. However, a combination of ISSR primers identified almost all the zygotic seedlings.