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Strand-specific RNA-seq is a powerful tool for the discovery of novel transcripts, annotation of genomes, and profiling of gene expression levels. Tn5 transposase has been successfully applied in massive-scale sequencing projects; in particular, Tn5 adaptor modification is used in epigenetics, genomic structure, and chromatin visualization. We developed a novel dU-adaptor-assembled Tn5-mediated strand-specific RNA-sequencing protocol and compared this method with the leading dUTP method in terms of experimental procedure and multiple quality metrics of the generated libraries. The results showed that the dU-Tn5 method is easy to operate and generates a strand-specific RNA-seq library of comparable quality considering library complexity, strand-specificity, evenness, and continuity of annotated transcript coverage. We also evaluated the performance of the dU-Tn5 method in identifying nitrogen-responsive protein-coding genes and long non-coding RNAs in soybean roots. The results indicated that ~62–70% of differentially expressed genes detected from conventional libraries were also detected in dU-Tn5 libraries, indicating good agreement of our method with the current standard; moreover, their fold-changes were highly correlated (R >0.9). Thus, our method provides a promising 'do-it-yourself' stranded RNA-seq procedure for gene expression profiling. [ABSTRACT FROM AUTHOR]

P2-type Ca2+ ATPases are responsible for cellular Ca2+ transport, which plays an important role in plant development and tolerance to biotic and abiotic stresses. However, the role of P2-type Ca2+ ATPases in stress response and stomatal regulation is still elusive in soybean. In this study, a total of 12 P2-type Ca2+ ATPases genes (GmACAs and GmECAs) were identified from the genome of Glycine max. We analyzed the evolutionary relationship, conserved motif, functional domain, gene structure and location, and promoter elements of the family. Chlorophyll fluorescence imaging analysis showed that vegetable soybean leaves are damaged to different extents under salt, drought, cold, and shade stresses. Real-time quantitative PCR (RT-qPCR) analysis demonstrated that most of the GmACAs and GmECAs are up-regulated after drought, cold, and NaCl treatment, but are down-regulated after shading stress. Microscopic observation showed that different stresses caused significant stomatal closure. Spatial location and temporal expression analysis suggested that GmACA8, GmACA9, GmACA10, GmACA12, GmACA13 , and GmACA11 might promote stomatal closure under drought, cold, and salt stress. GmECA1 might regulate stomatal closure in shading stress. GmACA1 and GmECA3 might have a negative function on cold stress. The results laid an important foundation for further study on the function of P2-type Ca2+ ATPase genes GmACAs and GmECAs for breeding abiotic stress-tolerant vegetable soybean. [ABSTRACT FROM AUTHOR]

Vegetable soybean is a type of value-added specialty soybean, served as a fresh vegetable or snack in China. Due to the difference from other types, it is important to understand the genetic structure and diversity of vegetable soybean for further utilization in breeding programs. The four vegetable cultivars, Taiwan-75, Zhexiandou No. 8, Zhexian No. 9 and Zhexian No. 10 are popular soybean varieties planted in Zhejiang province, and have large pods and intermediate maturity. The clustering showed a close relationship of these four cultivars in simple sequence repeat analysis. To reveal the genome variation of vegetable soybean, these four improved lines were analyzed by whole-genome re-sequencing. The average sequencing depth was 7X and the coverage ratio of each cultivar was at least more than 94%. Compared with the reference genome, a large number of single-nucleotide polymorphisms, insertion/deletions and structure variations were identified with different chromosome distributions. The average heterozygosity rate of the single-nucleotide polymorphisms was 11.99% of these four cultivars. According to the enrichment analysis, there were 23,371 genes identified with putative modifications, and a total of 282 genes were related to carbohydrate metabolic processes. These results provide useful information for genetic research and future breeding, which can facilitate the selection procedures in vegetable soybean breeding. [ABSTRACT FROM AUTHOR]

P 2 -type Ca 2+ ATPases are responsible for cellular Ca 2+ transport, which plays an important role in plant development and tolerance to biotic and abiotic stresses. However, the role of P 2 -type Ca 2+ ATPases in stress response and stomatal regulation is still elusive in soybean. In this study, a total of 12 P 2 -type Ca 2+ ATPases genes ( GmACAs and GmECAs ) were identified from the genome of Glycine max . We analyzed the evolutionary relationship, conserved motif, functional domain, gene structure and location, and promoter elements of the family. Chlorophyll fluorescence imaging analysis showed that vegetable soybean leaves are damaged to different extents under salt, drought, cold, and shade stresses. Real-time quantitative PCR (RT-qPCR) analysis demonstrated that most of the GmACAs and GmECAs are up-regulated after drought, cold, and NaCl treatment, but are down-regulated after shading stress. Microscopic observation showed that different stresses caused significant stomatal closure. Spatial location and temporal expression analysis suggested that GmACA8, GmACA9, GmACA10, GmACA12, GmACA13 , and GmACA11 might promote stomatal closure under drought, cold, and salt stress. GmECA1 might regulate stomatal closure in shading stress. GmACA1 and GmECA3 might have a negative function on cold stress. The results laid an important foundation for further study on the function of P 2 -type Ca 2+ ATPase genes GmACAs and GmECAs for breeding abiotic stress-tolerant vegetable soybean., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Wang, Fu, Zhang, Chen, Li, Shangguan, Zheng, Xu, Chen and Xu.)