Probabilistic alignment leads to improved accuracy and read coverage for bisulfite sequencing data.

BACKGROUND: DNA methylation has been linked to many important biological phenomena. Researchers have recently begun to sequence bisulfite treated DNA to determine its pattern of methylation. However, sequencing reads from bisulfite-converted DNA can vary significantly from the reference genome because of incomplete bisulfite conversion, genome variation, sequencing errors, and poor quality bases. Therefore, it is often difficult to align reads to the correct locations in the reference genome. Furthermore, bisulfite sequencing experiments have the additional complexity of having to estimate the DNA methylation levels within the sample. RESULTS: Here, we present a highly accurate probabilistic algorithm, which is an extension of the Genomic Next-generation Universal MAPper to accommodate bisulfite sequencing data (GNUMAP-bs), that addresses the computational problems associated with aligning bisulfite sequencing data to a reference genome. GNUMAP-bs integrates uncertainty from read and mapping qualities to help resolve the difference between poor quality bases and the ambiguity inherent in bisulfite conversion. We tested GNUMAP-bs and other commonly-used bisulfite alignment methods using both simulated and real bisulfite reads and found that GNUMAP-bs and other dynamic programming methods were more accurate than the more heuristic methods. CONCLUSIONS: The GNUMAP-bs aligner is a highly accurate alignment approach for processing the data from bisulfite sequencing experiments. The GNUMAP-bs algorithm is freely available for download at: http://dna.cs.byu.edu/gnumap. The software runs on multiple threads and multiple processors to increase the alignment speed.

Rhizobial plasmids that cause impaired symbiotic nitrogen fixation and enhanced host invasion.

The genetic rules that dictate legume-rhizobium compatibility have been investigated for decades, but the causes of incompatibility occurring at late stages of the nodulation process are not well understood. An evaluation of naturally diverse legume (genus Medicago) and rhizobium (genus Sinorhizobium) isolates has revealed numerous instances in which Sinorhizobium strains induce and occupy nodules that are only minimally beneficial to certain Medicago hosts. Using these ineffective strain-host pairs, we identified gain-of-compatibility (GOC) rhizobial variants. We show that GOC variants arise by loss of specific large accessory plasmids, which we call HR plasmids due to their effect on symbiotic host range. Transfer of HR plasmids to a symbiotically effective rhizobium strain can convert it to incompatibility, indicating that HR plasmids can act autonomously in diverse strain backgrounds. We provide evidence that HR plasmids may encode machinery for their horizontal transfer. On hosts in which HR plasmids impair N fixation, the plasmids also enhance competitiveness for nodule occupancy, showing that naturally occurring, transferrable accessory genes can convert beneficial rhizobia to a more exploitative lifestyle. This observation raises important questions about agricultural management, the ecological stability of mutualisms, and the genetic factors that distinguish beneficial symbionts from parasites.