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Isobaric labeling-based proteomics is widely applied in deep proteome quantification. Among the platforms for isobaric labeled proteomic data analysis, the commercial software Proteome Discoverer (PD) is widely used, incorporating the search engine CHIMERYS, while FragPipe (FP) is relatively new, free for noncommercial purposes, and integrates the engine MSFragger. Here, we compared PD and FP over three public proteomic data sets labeled using 6plex, 10plex, and 16plex tandem mass tags. Our results showed the protein abundances generated by the two software are highly correlated. PD quantified more proteins (10.02%, 15.44%, 8.19%) than FP with comparable NA ratios (0.00% vs. 0.00%, 0.85% vs. 0.38%, and 11.74% vs. 10.52%) in the three data sets. Using the 16plex data set, PD and FP outputs showed high consistency in quantifying technical replicates, batch effects, and functional enrichment in differentially expressed proteins. However, FP saved 93.93%, 96.65%, and 96.41% of processing time compared to PD for analyzing the three data sets, respectively. In conclusion, while PD is a well-maintained commercial software integrating various additional functions and can quantify more proteins, FP is freely available and achieves similar output with a shorter computational time. Our results will guide users in choosing the most suitable quantification software for their needs.

Scanning SWATH coupled with normal-flow LC has been recently introduced for high-content, high-throughput proteomics analysis, which requires a relatively large amount of sample injection. Here we established the microflow LC coupled with Scanning SWATH for samples with relatively small quantities. First, we optimized several key parameters of the LC and MS settings, including C18 particle size for the analytical column, LC gradient and flow rate, as well as effective ion accumulation time and isolation window width for MS acquisition. We then compared the optimized Scanning SWATH method with the conventional variable window SWATH (referred to as SWATH) method. Results showed that the total ion chromatogram signals in Scanning SWATH were 10 times higher than that of SWATH, and Scanning SWATH identified 12.2-22.2% more peptides than SWATH. Finally, we employed 120 min Scanning SWATH to acquire the proteomes of 62 formalin-fixed, paraffin-embedded (FFPE) tissue samples from 31 patients with hepatocellular carcinoma (HCC). Altogether, 92 334 peptides and 8516 proteins were quantified. Besides the reported biomarkers, including ANXA2, MCM7, SUOX, and AKR1B10, we identified new potential HCC biomarkers such as CST5, TP53, CEBPB, and E2F4. Taken together, we present an optimal workflow integrating microflow LC and Scanning SWATH that effectively improves the protein identification and quantitation.

An in vitro plant regeneration protocol of Cymbidium faberi from immature seeds was established. The immature seeds of 50 days old started to form rhizomes 4 months after they were cultured on hormone free medium. The rhizomes multiplied 5 times when subcultured on the medium containing 1.0 mg l−1α-naphthalene acetic acid (NAA) for 40 days and more than 90% of the rhizomes initiated shoots within 60 days on the media containing 0.5 or 1.0 mg l−1 NAA plus 2.0 or 5.0 mg l−1N6-benzylaminopurine (BA). Plantlets were regenerated when the shoots were planted on the basal medium amended with 1 g l−1 activated charcoal for 50 days and the plantlets grew normally after transplanting.

The development of the leaf sheaths in two species of Festuca, subgenus Festuca, Festuca trachyphylla (Hackel) Krajina with open leaf sheaths and Festuca rubra L. s.l. with closed leaf sheaths is compared to determine the developmental basis of this taxonomically important character. In both species serial cross sections of seedlings, scanning electron microscopy, and dissections of whole plants showed that a short tubular leaf base is formed early during the second plastochron of leaf development. Later during the same plastochron, ligule formation delimits sheath and blade; at this stage, the tubular portion occupies about one-half (200 – 500 μm) of the incipient leaf sheath. Differential extension growth of the distal open portion and proximal closed section of the sheath results in the primarily open or closed sheaths that characterize the mature leaves of the two species. The longitudinal splitting of the tubular leaf sheath in a position opposite the midvein in F. rubra is anticipated by a zone of small-diameter cells within the ground tissue, but no other structural correlates of sheath mechanical strength and persistence were detected. Our observations indicate that subtle shifts in the distribution of growth during leaf expansion result in the contrasting character states exemplified by these two species. Key words: Festuca, grass leaf development, leaf sheath, Poaceae.