Deletion mutagenesis identifies a haploinsufficient role for γ-zein in opaque2 endosperm modification.
Quality Protein Maize (QPM) is a hard kernel variant of the high-lysine mutant opaque2. Using γ-irradiation, we created opaque QPM variants to identify opaque2 modifier genes and to investigate deletion mutagenesis combined with Illumina sequencing as a maize (Zea mays) functional genomics tool. A K0326Y QPM deletion mutant was null for the 27- and 50-kD γ-zeins and abolished vitreous endosperm formation. Illumina exon and RNA sequencing revealed a 1.2-megabase pair deletion encompassing the 27- and 50-kD γ-zein genes on chromosome 7 and a deletion of at least 232 kb on chromosome 9. Protein body number was reduced by over 90%, while protein body size is similar to the wild type. Kernels hemizygous for the γ-zein deletion had intermediate 27- and 50-kD γ-zein levels and were semivitreous, indicating haploinsufficiency of these gene products in opaque2 endosperm modification. The γ-zein deletion further increased lysine in QPM in its homozygous and hemizygous states. This work identifies 27-kD γ-zein as an opaque2 modifier gene within the largest QPM quantitative trait locus and may suggest the 50-kD γ-zein also contributes to this quantitative trait locus. It further demonstrates that genome-wide deletions in nonreference maize lines can be identified through a combination of assembly of Illumina reads against the B73 genome and integration of RNA sequencing data.
Center for Plant Science Innovation, Beadle Center for Biotechnology , University of Nebraska, Lincoln, Nebraska 68588.
Lingling
L
Yuan
Yongchao
Y
Dou
Shahryar F
SF
Kianian
Chi
C
Zhang
David R
DR
Holding
D032461:Chromosomes, Plant; D056625:Endosperm / Q000235:genetics*; D005091:Exons; D005720:Gamma Rays; D017353:Gene Deletion; D018506:Gene Expression Regulation, Plant; D057895:Haploinsufficiency / Q000235:genetics*; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D040641:Quantitative Trait Loci; D018749:RNA, Plant; D003313:Zea mays / Q000235:genetics* / Q000528:radiation effects; D015028:Zein / Q000235:genetics*
D010940:Plant Proteins; D018749:RNA, Plant; D015028:Zein; D008239:Lysine