Improving the sorting efficiency of maize haploid kernels using an NMR-based method with oil content double thresholds

BACKGROUND: Maize haploid breeding technology can be used to rapidly develop homozygous lines, significantly shorten the breeding cycle and improve breeding efficiency. Rapid and accurate sorting haploid kernels is a prerequisite for the large-scale application of this technology. At present, the automatic haploid sorting based on nuclear magnetic resonance (NMR) using a single threshold method has been realized. However, embryo-aborted (EmA) kernels are usually produced during in vivo haploid induction, and both haploids and EmA kernels have lower oil content and are separated together using a single threshold method based on NMR. This leads to a higher haploid false discrimination rate (FDR) and requires secondary manual sorting to select the haploid kernels from the mixtures, which increases the sorting cost and decreases the haploid sorting efficiency. In order to improve the correct discrimination rate (CDR) in sorting haploids, a method to distinguish EmA kernels is required. RESULTS: Single kernel weight and oil content were measured for the diploid, haploid, and EmA kernels derived from three maize hybrids and nine inbred lines by in vivo induction. The results showed that the distribution of oil content showed defined boundaries between the three types of kernels, while the single kernel weight didn't. According to the distribution of oil content in the three types of kernels, a double-threshold method was proposed to distinguish the embryo-aborted kernels, haploid and diploid kernels based on NMR and their oil content. The double thresholds were set based on the minimum oil content of diploid kernels and the maximum content of EmA kernels as the upper and lower boundary values, respectively. The CDR of EmA kernels in different maize materials was > 97.8%, and the average FDR was reduced by 27.9 percent. CONCLUSIONS: The oil content is an appropriate indicator to discriminate diploid, haploid and EmA kernels. An oil content double-threshold method based on NMR was first developed in this study to identify the three types of kernels. This methodology could reduce the FDR of haploids and improve the sorting efficiency of automated sorting system. Thus, this technique represents a potentially efficient method for haploid sorting and provides a reference for the process of automated sorting of haploid kernels with high efficiency using NMR.