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<p>Abstract</p> <p>Accurate analysis of DNA methylation by bisulphite sequencing depends on the complete conversion of all cytosines into uracil. Until now there has been no standard or universal gene identified as an endogenous control to monitor the conversion frequency in plants. Here, we report the development of PCR based assays for one nuclear gene <it>IND </it>(<it>INDEHISCENT</it>) and two mitochondrial genes, <it>NAD </it>(<it>NICOTINAMIDE ADENINE DINUCLEOTIDE</it>) and <it>ATP1 </it>(<it>ATPase SUBUNIT 1</it>). We demonstrated their efficacy as bisulphite conversion controls in <it>Brassica </it>and other plant taxa. The target regions amplified by four primer pairs were found to be consistently free from DNA methylation. Primer pairs for <it>IND.a </it>and <it>NAD </it>were effective within <it>Brassica </it>species, whereas two primer pairs for <it>ATP1 </it>provided reliable controls across a representative range of dicot and monocot angiosperm species. These primer sets may therefore be adopted as controls in plant methylation analysis for a wide range of studies.</p>