Effect of Sample Preparation Methods on the Quality of RNA Extraction from Chrysanthemum Bud Laser Microdissection Samples
[Objective]To study the effect of different fixatives and section methods on RNA quality of flower primordia of Chrysanthemum buds collected by microdissection.[Method]Chrysanthemum buds was fixed by acetone or ethanol acetic acid and then paraffin section/frozen section was prepared for laser microdissection respectively.Outermost two whorls of flower primordia were identified and collected by laser mi-crodissection.The quantity and quality of isolated RNA were assessed.[Result]Both tissues embedded in paraffin for paraffin section or OCT for cryosection preserves well the anatomical structure and morphology of the buds.For paraffin-section,ethanol acetic acid was a better choice be-ing a fixative than that of acetone,as that RNA extracted from the microdissection samples was less degraded.There was no significant difference in RNA integrity of cryosection between these two fixatives.The tissues conducted cryosection could get higher quality of RNA compared with that of paraffin-section.Furthermore,when microdissection was conducted,containing the extraction buffer from RNA isolation kit in the cap of PCR tube could preserved RNA integrity of the samples.Genes expression of flowering locus T in flower primordia was different in species of chrysanthemum showed that samples captured by microdissection can be used for subsequent research.[Conclusion]Plant tissues fixed by etha-nol-acetic acid and protected by gradient sucrose before cryosection could collect specific cell-type populations by laser microdissection. From those populations we could obtain enough RNA with high integrity.
万方数据
维普
