New methods for computational decomposition of whole-mount in situ images enable effective curation of a large, highly redundant collection of Xenopus images

The precise anatomical location of gene expression is an essential component of the study of gene function. For most model organisms this task is usually undertaken via visual inspection of gene expression images by interested researchers. Computational analysis of gene expression has been developed in several model organisms, notably in Drosophila which exhibits a uniform shape and outline in the early stages of development. Here we address the challenge of computational analysis of gene expression in Xenopus, where the range of developmental stages of interest encompasses a wide range of embryo size and shape. Embryos may have different orientation across images, and, in addition, embryos have a pigmented epidermis that can mask or confuse underlying gene expression. Here we report the development of a set of computational tools capable of processing large image sets with variable characteristics. These tools efficiently separate the Xenopus embryo from the background, separately identify both histochemically stained and naturally pigmented regions within the embryo, and can sort images from the same gene and developmental stage according to similarity of gene expression patterns without information about relative orientation. We tested these methods on a large, but highly redundant, collection of 33,289 in situ hybridization images, allowing us to select representative images of expression patterns at different embryo orientations. This has allowed us to put a much smaller subset of these images into the public domain in an effective manner. The ‘isimage’ module and the scripts developed are implemented in Python and freely available on https://pypi.python.org/pypi/isimage/.Author summary: An important component of research into the function of genes in the developing organism is an understanding of both when and where the gene is expressed. Well established molecular techniques can be used to colour the embryo in regions where the gene of interest appears, and researchers will photograph such treated embryos at different stages of development to build up the story of the gene’s use. Small numbers of these expression pattern images may easily be examined by eye, but getting usable information from large collections of such images would take an enormous investment in time by trained scientists. Computational analysis is much to be preferred, but the task is complex and difficult to generalise. The frog Xenopus is an important model for studying vertebrate development, but up till now has had no purely computational methods available for analysing gene expression. Here we present a suite of computational tools based on a range of mathematical methods, capable of recognising the outline of the embryo against a variety of backgrounds, and within the embryo separately recognising areas of both gene expression and natural pigmentation. These tools work over a wide range of embryo shapes and imaging conditions, and, in our opinion, represent a major step towards full automation of anatomical gene expression annotation in vertebrate embryology.