Protocol for flow cytometric estimation of nuclear DNA amount in fig (Ficus carica L.) cultivars

R. Pio, A.C.L. de Oliveira, M. Pasqual, L.A.S. Pio, P.N. Curi, R.B. Bisi
The present study was undertaken to develop a protocol to differentiate fig cultivars by nuclear DNA content using flow cytometric estimation method. Eighteen fig cultivars were used in this study in a randomized design with nine repeations. We evaluated three buffers extraction protocol (LB01, Marie and Tris MgCl2), the effect of presence or absence of RNase, different doses of Propidium Iodide (5, 10, 15, 20, 25 and 30 µL of a solution of 1 mg mL-1 Propidium Iodide), different periods of exposure to Propidium Iodide (0, 5, 10, 15 and 20 min) and six reference standards (peas, fava beans, radish, rye, soybean and tomato) The coefficient of variation (CV) and DNA contents were determined Statistical analysis were made using the SISVAR® software. We observed the most appropriate buffer is Marie. The volume of Propidium Iodide is 10 µL without addition of RNase, and the exposure time of iodide in the sample of 5 min. The reference standard recommended is Pisum sativum L. The estimate of the DNA content obtained by flow cytometry enables to differentiate the fig cultivars.
Pio, R., de Oliveira, A.C.L., Pasqual, M., Pio, L.A.S., Curi, P.N. and Bisi, R.B. 2017. Protocol for flow cytometric estimation of nuclear DNA amount in fig (Ficus carica L.) cultivars. Acta Hort. (ISHS) 1173:99-104
http://www.actahort.org/books/1173/1173_17.htm
Ficus carica L., DNA content, flow cytometry, protocol
English
1173_17
99-104

Acta Horticulturae