Fine mapping of yll prioritizes aberrantly spliced Glyma.10G090700 (GmPAO7) as the strongest candidate in a soybean yellow-leaf lethal mutant

Abstract Chlorophyll-deficient mutants are useful for dissecting chloroplast biology in crops. Here, we characterized a recessive soybean ( Glycine max (L.) Merr.) yellow-leaf lethal mutant, yll , that showed markedly reduced chlorophyll content, impaired photosystem II performance, and seedling lethality. Fine mapping delimited yll to a 165-kb interval on chromosome 10. In the Wm82.a6.v1 annotation, this interval contains two genes, of which Glyma.10G090700 ( GmPAO7 ) was prioritized as the strongest candidate on the basis of leaf expression, reduced transcript abundance in yll , and a reproducible intron-retention event. Retention of the 110-bp intron between exons 1 and 2 introduced a premature termination codon and was predicted to shorten the annotated 500-aa PAO1-like protein to 32 aa, eliminating the annotated flavin amine oxidase and NAD(P)-binding domains. Direct sequencing of RT-PCR products detected the normally spliced transcript in wild-type plants and the intron-retained sequence in mutant plants, with overlapping splice forms in heterozygous plants; this qualitative assay does not establish splice-isoform proportions. RNA-seq identified broad expression differences between wild-type and mutant sibling seedlings, including enrichment of photosynthesis-, chloroplast-, and redox-associated categories; because yll seedlings were severely chlorotic and developmentally compromised, these profiles are interpreted as the transcriptomic state associated with the phenotype and may largely reflect downstream responses. Transmission electron microscopy revealed chloroplast ultrastructural abnormalities associated with yll , including poorly developed grana and disorganized thylakoid membranes, but these observations do not functionally validate the candidate gene. Together, the mapping and transcript evidence prioritize Glyma.10G090700 ( GmPAO7 ) as the strongest candidate for yll ; Glyma.10G091000 remains in the interval, and functional complementation or an independent allele will be required to establish causality. A PAO–polyamine–ROS connection remains a testable hypothesis because PAO activity, polyamine concentrations, and ROS were not measured.