Establishment and Optimization of an Experiment System for Flow Cytometry in Oil-Seed Camellia

Abstract

Ploidy identification is a basic method for determining germplasm resources and for breeding new varieties of oil-seed camellia. In this study, flow cytometry and K-mer analysis were used to identify the ploidy of oil-seed camellia germplasms. To determine the best tissue organ type, lysis time, and dyeing time, evaluation indices such as the presence of a clear main peak, the ease of sampling, and the coefficient of variation were used. A technique was established, and the ploidies of different oil-seed camellia germplasms were identified. The results showed that pollen was the best material and that using a 400 mL PI lysis solution for 10 min lysis, followed by dyeing with a 1600 mL DAPI dyeing solution for 10 min, was the most suitable technique. According to the peak value of the control diploid Camellia azalea, 15 samples were estimated to be diploid, 24 samples were tetraploid, 18 samples were hexaploid, and 13 samples were octoploid. In addition, the K-mer analysis results showed that among the five samples, CK, C051, and C047 were diploid, while C037 and C043 were tetraploid, results that are consistent with the results of the flow cytometry identification. This study is therefore valuable for the polyploid selection and use of different ploidy germplasm resources for the cross breeding of oil-seed camellia.

Keywords

• oil-seed camellia
• germplasm resources
• ploidy
• flow cytometry
• diploid