Identification of self-incompatibility in macadamia (Macadamia SPP.) using field-bagging and fluorescence microscopy

Self-incompatibility (SI) significantly reduces crop yield, often far below its genetic potential. Developing self-compatible varieties is the most effective strategy for overcoming SI in crops. Most macadamia (Macadamia SPP.) species exhibit SI or partial self-incompatibility (PSI), so the efficient identification of self-compatible germplasms has emerged as a crucial topic. To characterize self-incompatibility phenotypes in macadamia germplasm resources, we conducted a four-year field study using the field-bagging method and established a standardized classification system of self-incompatibility. That is, the degree of SI was based on the final self-incompatibility index (F_SI), which was calculated based on the open-pollination final nut set per raceme (OP_FNS) and self-pollination final nut set per raceme (SP_FNS) values (strong SI: F_SI ≥ 0.7, medium SI: 0.4 ≤ F_SI <0.7, and weak SI: F_SI < 0.4). To develop a method for identifying macadamia SI more efficient and cost-effective than the field-bagging method, we employed pollen tube fluorescence staining, where the degree of SI was classified based on the percentage of pistils for which the pollen tube length exceeded 7/10 of the style (PLS) (strong SI: PLS ≤ 16%, medium SI: 16% < PLS ≤ 35%, and weak SI: PLS > 35%). Through comprehensive analysis of the field-bagging and fluorescence-microscopy observations, thirteen varieties with strong SI (816, 778, 842, Special, 812, D, 820, 246, 772, A16, A4, 951, and 695), six varieties with moderate SI (851, 828, 508, 936, O.C, and D4), and four varieties with weak SI (915, HY, 836, and 814) were identified. Our study provides a theoretical foundation and technical support for advancing germplasm resource innovation, and the genetic improvement and breeding of self-compatible macadamia varieties.