Double fertilization is a characteristic reproductive style in flowering plants. Two immobile sperm cells are delivered into the ovule by a pollen tube and then fertilize with two female gametes (egg cell and central cell). Several sperm cell-specific membrane proteins localizing on the cell surface have been characterized to control fertilization (Fertilization factors). The functions of the reported fertilization factors have been mainly characterized using T-DNA insertion, single nucleotide substitution, or partially edited mutant Arabidopsis lines. However, such mutants might have the possibility to affect the functional analyses due to unpredictable and abnormal protein expression from the remaining coding region. In this study, we generated novel mutants lacking an open reading frame (ORF) of fertilization factors to reanalyze their functions precisely. CRISPR/Cas9 technology was applied using multiple guide RNAs, which target the region close to the start and stop codons. The deletions of the ORFs in T1 plants were confirmed by PCR and sequencing, showing the edition efficiency was 75% (15/20). T1 mutant lines lacking ORF showed undeveloped seed phenotype, as the mark of fertilization failure observed in the previously reported T-DNA mutant line. Additionally, we obtained mutant lines which produce aborted seeds, the phenotype caused by single fertilization of central cell. This is the novel phenotype never observed before in the T-DNA insertion mutant. In this presentation, we will report the progress of the investigation of the novel type of mutant.