Main conclusion <p>This study uncovered the genetic basis of leaf variegation for 35 diverse flowering plants. The underlying loss-of-function plastome mutations in albino sectors include single nucleotide substitutions and insertions/deletions in protein genes mostly related to transcription and photosynthesis.</p> Abstract <p>Variegated plants have high ornamental value in horticulture. However, the genetic basis of plant variegation has not been systematically investigated. Here we investigated the genetic basis of variegation for 46 plants by comparing the plastid genomes (plastomes) of green and albino sectors of each plant. Our analyses revealed that 35 plants each had a single plastome mutation in their albino sectors, while the remaining 11 had no plastome mutations, which may be caused by mutations in their nuclear or mitochondrial genomes. These 35 mutations include 12 single nucleotide substitutions, and 17 small (1–10 bp), 3 medium (11–100 bp) and 3 large (&gt; 100 bp) insertions/deletions (indels) in protein genes mostly related to transcription and photosynthesis. All the single nucleotide substitutions cause premature stop codons, 19 of the 20 small and medium indels generate frameshifts and premature stop codons, and all three large indels eliminate multiple genes. The remaining one is a 6-bp deletion occurring in the intron of <i>rps16</i>, which may cause a splicing defect. Dysfunction of these genes due to mutations can explain leaf variegation in the 35 plants. Combining with previous studies, we found that plastome mutations leading to leaf variegation occur predominantly in the coding regions of the LSC and much more common in <i>rpo</i> genes than other genes. Relative to single nucleotide substitutions, indels are much more common. Over half of&#xa0;the detected indels are copy-number variations, suggesting replication slippage is a major mechanism of plastome mutations. Our study deepens the understanding of the genetic basis of plant variegation.</p>