Leaf senescence constitutes a highly regulated final phase of leaf development, leading to cell death that is recognized as a type of programmed cell death (PCD). Degradation of nuclear DNA into oligonucleosomal fragments (DNA ladder) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay are methods commonly used to detect PCD-specific DNA cleavage. TUNEL reaction in situ labels free 3′-OH DNA strand breaks (nicks), thus allowing histological localization of nuclear DNA degradation during PCD. Here we describe in situ labeling of PCD-specific nuclear DNA fragmentation on conventional histological sections of senescing tobacco leaves. Incorporation of fluorescein-labeled dUTPs is detected by fluorescence microscopy, which enables in situ visualization of PCD at the single-cell level in the leaf mesophyll tissues undergoing senescence.
CITATION STYLE
Uzelac, B., Janošević, D., & Budimir, S. (2018). In situ detection of programmed cell death in senescing nicotiana tabacum leaves using TUNEL assay. In Methods in Molecular Biology (Vol. 1744, pp. 267–282). Humana Press Inc. https://doi.org/10.1007/978-1-4939-7672-0_21
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