Motivation: Bulked segregant analysis combined with next generation sequencing has proven to be a simple and efficient approach for fast mapping of quantitative trait loci (QTLs). However, how to estimate the proportion of phenotypic variance explained by a QTL (or termed QTL heritability) in such pooled QTL mapping is an unsolved problem. Results: In this paper, we propose a method called PQHE to estimate QTL heritability using pooled sequencing data obtained under different experimental designs. Simulation studies indicated that our method is correct and feasible. Four practical examples from rice and yeast are demonstrated, each representing a different situation. Availability and implementation: The R scripts of our method are open source under GPLv3 license at http://genetics.fafu.edu.cn/PQHE or https://github.com/biotangweiqi/PQHE. The R scripts require the R package rootSolve. Contact: [email protected]. Supplementary information: Supplementary data are available at Bioinformatics online.