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Updated: May 23, 2026

Preparation of Intact Tissue for Microscopic Analysis of the Endosperm Cell Layer in Developing and Mature Arabidopsis Seeds
Published on: May 16, 2025
Arabidopsis ENDO2: its catalytic role and requirement of N-glycosylation for function
Chia-Yun Ko1, Yi-Ling Lai, Wen-Yu Liu
1Institute of Bioinformatics and Structural Biology, National Tsing Hua University , Hsinchu, 30013, Taiwan.
Abstract:
The Arabidopsis thaliana At1g68290 gene encoding an endonuclease was isolated and designated ENDO2, which was cloned into a binary vector to overexpress ENDO2 with a C-terminal 6 × His-tag in A. thaliana. Our Arabidopsis transgenic lines harboring 35SP::ENDO2 produced stable active enzyme with high yield. The protein was affinity purified from transgenic plants, and its identity was confirmed by liquid chromatography-mass spectrometry and automatic Edman degradation. ENDO2 enzyme digests RNA, ssDNA, and dsDNA, with a substrate preference for ssDNA and RNA. The activity toward ssDNA (361.7 U/mg) is greater than its dsDNase activity (14.1 U/mg) at neutral pH. ENDO2 effectively cleaves mismatch regions in heteroduplex DNA containing single base pair mismatches or insertion/deletion bases and can be applied to high-throughput detection of single base mutation. Our data also validated that the removal of sugar groups from ENDO2 strongly affects its enzymatic stability and activity.
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