Genome assembly of the Chinese maize elite inbred line RP125 and its EMS mutant collection provide new resources for maize genetics research and crop improvement

Abstract

SUMMARY Maize is an important crop worldwide, as well as a valuable model with vast genetic diversity. Accurate genome and annotation information for a wide range of inbred lines would provide valuable resources for crop improvement and pan‐genome characterization. In this study, we generated a high‐quality de novo genome assembly (contig N50 of 15.43 Mb) of the Chinese elite inbred line RP125 using Nanopore long‐read sequencing and Hi‐C scaffolding, which yield highly contiguous, chromosome‐length scaffolds. Global comparison of the RP125 genome with those of B73, W22, and Mo17 revealed a large number of structural variations. To create new germplasm for maize research and crop improvement, we carried out an EMS mutagenesis screen on RP125. In total, we obtained 5818 independent M2 families, with 946 mutants showing heritable phenotypes. Taking advantage of the high‐quality RP125 genome, we successfully cloned 10 mutants from the EMS library, including the novel kernel mutant qk1 (quekou: “missing a small part” in Chinese), which exhibited partial loss of endosperm and a starch accumulation defect. QK1 encodes a predicted metal tolerance protein, which is specifically required for Fe transport. Increased accumulation of Fe and reactive oxygen species as well as ferroptosis‐like cell death were detected in qk1 endosperm. Our study provides the community with a high‐quality genome sequence and a large collection of mutant germplasm.

A telomere-to-telomere gap-free reference genome of watermelon and its mutation library provide important resources for gene discovery and breeding

“…Using pollen-EMS mutagenesis, the watermelon phenotypic mutation frequency (15.38%) was similar to that reported in the maize inbred RP125 mutant library (16.26%) (Nie et al, 2021) but much higher than that for the tropical maize inbred ML10 mutant library (6.16%) (Tran et al, 2020). The gap-free genome assembled in this study facilitated our identification and characterization of genome-wide mutations.…”

“…A high-quality genome combined with corresponding large mutant collections will facilitate the identification and cloning of genes. This strategy has been used to clone many genes from EMS libraries in maize (Nie et al, 2021) and rapeseed (Tang et al, 2020). In our study, taking advantage of the gap-free G42 genome, a dominant malesterility gene ClMS1 was quickly identified from the EMS mutant library by whole-genome re-sequencing of only 23 individual plants with an average sequence depth of 57.23.…”

“…An alternative approach introduced and used extensively in maize is to use EMS-mutagenized pollen grains for artificial pollination (Neuffer, 1994). Soaking pollen grains in a suspension of EMS in inert mineral oil has been shown to create a large number of mutants, and several maize mutagenesis populations have been generated (Lu et al, 2018;Heuermann et al, 2019;Tran et al, 2020;Nie et al, 2021). Despite having higher mutation rates and a shorter screening time, this method has not yet been examined in other crops.…”

Identification and Fine Mapping of the Recessive Gene BK-5, Which Affects Cell Wall Biosynthesis and Plant Brittleness in Maize

“…Using these pairs of SSR markers and 94 B73 × bk-5 F2 individuals, we preliminarily mapped bk-5 between the flanking markers SSR11 and Indel-33 on chromosome 3 to a region about 16.3 Mb long ( Figure 4 a, Supplementary Table S2 ). Based on the published RP125 genome [ 52 ], more SSR/indel molecular markers were developed and an additional 412 mutants were also used to fine-map the mutation to a 90.2-Kb region ( Figure 4 a, Supplementary Table S3 ). Based on the reference genome, three putative open reading frames (ORFs) were located within this region ( Figure 4 a).…”

“…The pollen of RP125 was treated with a 0.1% solution of EMS (Sigma-Aldrich, St. Louis, MO, USA, M0880), dissolved in mineral oil (Sigma-Aldrich, St. Louis, MO, USA, M8410) for 30–40 min, then used to pollinate 60–70 ears to induce mutation. With this method, the EMS mutant library of the RP125 background was constructed by our research group [ 52 ]. We screened a series of mutants related to brittle stalk mutation in the library, and obtained a stable genetic brittle stalk mutant, named bk-5, through continuous selfing screening.…”