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A Mouse Model for Corneal Neovascularization by Alkali Burn
Published on: June 30, 2023
RNA Isolation and qPCR Analysis from Rat Corneal Tissue Following Alkali Burn Injury
1Department of Ophthalmology and Visual Science, Yeouido St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Republic of Korea.
Purpose:
To establish a practical protocol for RNA isolation and quantitative real-time polymerase chain reaction (qPCR) analysis using rat corneal tissue following alkali burn injury.
Methods:
Corneal alkali burn injury was induced in rats using a 4-mm filter, paper soaked in 1N NaOH applied to the central cornea for 40 seconds, followed by saline irrigation. Corneal tissues were collected 7 days after injury, and total RNA was isolated using a TRIzol-based protocol optimized for individual corneal samples. RNA concentration and purity were evaluated using spectrophotometry and agarose gel electrophoresis. Complementary DNA was synthesized for qPCR analysis of inflammation- and angiogenesis-related genes.
Results:
The optimized protocol enabled successful RNA isolation from individual rat corneal tissues without sample pooling. RNA quality was suitable for downstream qPCR analysis. Expression levels of Vegfa, Mmp9, and Il-6 were successfully detected and showed expected alterations in alkali-burned corneas.
Conclusion:
This protocol provides a practical and reproducible method for RNA isolation and qPCR analysis in rat corneal alkali burn models and may facilitate molecular investigations of ocular surface inflammation and wound healing.
