MethPrimer is a program designed for designing primers for bisulfite-conversion-based methylation PCRs, such as bisulfite sequencing PCR (BSP) and methylation-specific PCR (MSP). It is based on Primer3 and integrates CpG island prediction into primer design. MethPrimer can design primers for BSP and MSP, and also for other methods like COBRA and MS-SNuPE with minor modifications. The program takes a DNA sequence as input and searches for potential CpG islands. Primers are then selected around these islands or regions specified by the user. MethPrimer delivers results through a web browser in text and graphic view. CpG islands are defined as regions of DNA greater than 200 bp with a guanine/cytosine content above 0.5 and an observed or expected presence of CpG above 0.6. MethPrimer uses a sliding window algorithm to predict CpG islands. It then applies specific rules for primer selection based on the size of the CpG island and the desired product size. For BSP, primers should not contain any CpG sites to avoid discrimination between methylated and unmethylated DNA. For MSP, primers should contain at least one CpG site, preferably at the 3'-end. MethPrimer also ensures that primers for M and U pairs contain the same CpG sites. The program also considers the melting temperature (Tm) of primers and ensures that the Tm values of the two primer pairs are similar. MethPrimer was developed on a Linux platform and is accessible via a web browser. It was tested with random DNA sequences ranging from 10 to 100 kb in length, and its performance was found to be linear with respect to the length of the query sequence. MethPrimer is optimized for bisulfite-conversion-based PCRs and is designed to handle the specific constraints of these methods, such as the need for longer primers and the requirement for a certain number of 'C's in the primer sequence to distinguish between modified and unmodified DNA. The program is freely accessible at http://itsa.ucsf.edu/~urolab/methprimer.MethPrimer is a program designed for designing primers for bisulfite-conversion-based methylation PCRs, such as bisulfite sequencing PCR (BSP) and methylation-specific PCR (MSP). It is based on Primer3 and integrates CpG island prediction into primer design. MethPrimer can design primers for BSP and MSP, and also for other methods like COBRA and MS-SNuPE with minor modifications. The program takes a DNA sequence as input and searches for potential CpG islands. Primers are then selected around these islands or regions specified by the user. MethPrimer delivers results through a web browser in text and graphic view. CpG islands are defined as regions of DNA greater than 200 bp with a guanine/cytosine content above 0.5 and an observed or expected presence of CpG above 0.6. MethPrimer uses a sliding window algorithm to predict CpG islands. It then applies specific rules for primer selection based on the size of the CpG island and the desired product size. For BSP, primers should not contain any CpG sites to avoid discrimination between methylated and unmethylated DNA. For MSP, primers should contain at least one CpG site, preferably at the 3'-end. MethPrimer also ensures that primers for M and U pairs contain the same CpG sites. The program also considers the melting temperature (Tm) of primers and ensures that the Tm values of the two primer pairs are similar. MethPrimer was developed on a Linux platform and is accessible via a web browser. It was tested with random DNA sequences ranging from 10 to 100 kb in length, and its performance was found to be linear with respect to the length of the query sequence. MethPrimer is optimized for bisulfite-conversion-based PCRs and is designed to handle the specific constraints of these methods, such as the need for longer primers and the requirement for a certain number of 'C's in the primer sequence to distinguish between modified and unmodified DNA. The program is freely accessible at http://itsa.ucsf.edu/~urolab/methprimer.