A comparison of the actions of trypsin and pepsin on porcine immunoglobulin M and their effects on biological activity

The action of trypsin at 55 degree C and pH 8.3 on pig IgM anti-Salmonella has been compared with the action of pepsin at 37 degree C and pH 4.6. Both processes cause the gradual removal of Fab arms and Cmu2 domains to produce eventually an (Fc)5 fragment. However, during tryptic digestion Fab arms are preferentially removed from the same subunit, whereas peptic digestion causes random removal from any subunit. At intermediate stages of digestion both processes produce partially fragmented molecules which consist of an (Fc)5 portion still attached to limited numbers of Fab arms. Both processes cause a gradual decrease in the ability of molecules to agglutinate Salmonella, but complement fixation by the complexes declines much more rapidly. A stage is reached where molecules having four Fab arms can still agglutinate but there is no complement fixation. However, the remaining arms on the tryptic molecules are distributed in pairs on the same subunit, whereas those on the peptic molecules are distributed randomly. Hence the number of remaining Fab arms, rather than their distribution, appears to be the critical factor which influences biological activity. A possible explanation for this is discussed.     

Illuminating Choices for Library Prep: A Comparison of Library Preparation Methods for Whole Genome Sequencing of Cryptococcus neoformans Using Illumina HiSeq

The industry of next-generation sequencing is constantly evolving, with novel library preparation methods and new sequencing machines being released by the major sequencing technology companies annually. The Illumina TruSeq v2 library preparation method was the most widely used kit and the market leader; however, it has now been discontinued, and in 2013 was replaced by the TruSeq Nano and TruSeq PCR-free methods, leaving a gap in knowledge regarding which is the most appropriate library preparation method to use. Here, we used isolates from the pathogenic fungi Cryptococcus neoformans var. grubii and sequenced them using the existing TruSeq DNA v2 kit (Illumina), along with two new kits: the TruSeq Nano DNA kit (Illumina) and the NEBNext Ultra DNA kit (New England Biolabs) to provide a comparison. Compared to the original TruSeq DNA v2 kit, both newer kits gave equivalent or better sequencing data, with increased coverage. When comparing the two newer kits, we found little difference in cost and workflow, with the NEBNext Ultra both slightly cheaper and faster than the TruSeq Nano. However, the quality of data generated using the TruSeq Nano DNA kit was superior due to higher coverage at regions of low GC content, and more SNPs identified. Researchers should therefore evaluate their resources and the type of application (and hence data quality) being considered when ultimately deciding on which library prep method to use.     

Future directions in plant hormone research

Future directions in plant hormone research are discussed, with particular reference to the regulation of hormone biosynthesis, hormone perception, and signal transduction.