Thermal degradation of citrus pectin in low-moisture environment – Investigation of backbone depolymerisation

Thermal degradation of modified pectin samples with varying molecular structure during storage was recently studied at 60 °C and 80% relative humidity (rh) for 28 days. Demethoxylation and depolymerisation were identified as main degradation reactions. The present paper aims on improving the understanding of the different depolymerisation reactions and their interplay with demethoxylation during storage. Therefore, thermal degradaton of acidic and alkaline demethoxylated pectins was studied at a further reduced rh of 40%. The alterations were examined in detail via molecular parameters and were reflected by differential scanning calorimetry and attenuated total reflectance Fourier-transformation infrared spectroscopy. The impact of thermal degradation on pectin particle structure was studied via particle surface area and microscopy. At low relative humidity (rh) demethoxylation and depolymerisation were reduced, and the formation of brown reaction products, resulting from further decomposition of intermediate uronides and neutral sugars, was restricted. By comparing thermal degradation at different humidity, eliminative decarboxylation was identified as the main depolymerisation reaction. Reduction of rh affected also the alteration of pectin material properties, particle surface reduction was less pronounced. Molecular alterations were stronger in case of acidic demethoxylated samples, and alterations of material properties were higher in case of alkaline demethoxylated samples.DFG, 268547215, Strukturabhängige Abbaureaktionen von Pektinen und deren Auswirkungen auf nicht-enzymatische Bräunung und technologische Funktionalitä

A high-resolution single-molecule sequencing-based Arabidopsis transcriptome using novel methods of Iso-seq analysis

Background: Accurate and comprehensive annotation of transcript sequences is essential for transcript quantification and differential gene and transcript expression analysis. Single-molecule long-read sequencing technologies provide improved integrity of transcript structures including alternative splicing, and transcription start and polyadenylation sites. However, accuracy is significantly affected by sequencing errors, mRNA degradation, or incomplete cDNA synthesis. Results: We present a new and comprehensive Arabidopsis thaliana Reference Transcript Dataset 3 (AtRTD3). AtRTD3 contains over 169,000 transcripts—twice that of the best current Arabidopsis transcriptome and including over 1500 novel genes. Seventy-eight percent of transcripts are from Iso-seq with accurately defined splice junctions and transcription start and end sites. We develop novel methods to determine splice junctions and transcription start and end sites accurately. Mismatch profiles around splice junctions provide a powerful feature to distinguish correct splice junctions and remove false splice junctions. Stratified approaches identify high-confidence transcription start and end sites and remove fragmentary transcripts due to degradation. AtRTD3 is a major improvement over existing transcriptomes as demonstrated by analysis of an Arabidopsis cold response RNA-seq time-series. AtRTD3 provides higher resolution of transcript expression profiling and identifies cold-induced differential transcription start and polyadenylation site usage. Conclusions: AtRTD3 is the most comprehensive Arabidopsis transcriptome currently. It improves the precision of differential gene and transcript expression, differential alternative splicing, and transcription start/end site usage analysis from RNA-seq data. The novel methods for identifying accurate splice junctions and transcription start/end sites are widely applicable and will improve single-molecule sequencing analysis from any species