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Abstract [Objective] In order to optimize the efficient virus elimination and regeneration system from shoot apical meristem of tea chrysanthemum, suitable heat treatment duration and detoxification agents, this study aims to improve the problem of low virus-free seedlings obtaining ratio. [Methods] Six tea chrysanthemum varieties were used as materials. The axenic seedlings were pre-treated with alternating heat temperature treatment of 40 ℃/32 ℃ day/night, detoxification agents such as salicylic acid, ribavirin, and abscisic acid, as well as a combination of detoxification agents and heat treatment. The rate of callus induction, regeneration frequency, and average number of regenerated shoots from inoculated shoot apical meristems were compared among different treatment methods, the detoxification efficacy of various treatments were evaluated. [Results] There was a significant difference in the regeneration frequency of shoot apical meristems among different varieties of tea chrysanthemum, with ‘Wuyuan Huangju’ having the highest regeneration frequency of 54.77%. Under different heat treatment days, the difference in regeneration of shoot apical meristems in tea chrysanthemum reached a significant level, with ‘Hangbaiju’ and ‘Qiyuexue’ showing the best regeneration after 40 ℃/32 ℃ heat treatment. Salicylic acid, ribavirin, and abscisic acid treatments inhibited the regeneration of shoot apical meristems. Salicylic acid and ribavirin treatments combined with heat treatment promoted the regeneration of shoot apical meristems, while high concentration abscisic acid combined with heat treatment severely inhibited the regeneration of shoot apical meristems. Treatment with 10 μmol/L salicylic acid combined with heat treatment showed the best regeneration. Virus-free seedlings were obtained through suitable heat treatment combined with detoxification agents. [Conclusion] The combination of 10 μmol/L salicylic acid with 40 ℃/32 ℃ day/night pretreatment for 40 days significantly enhances the regeneration frequency of shoot apical meristems, and obtains virus-free seedlings

Sorghum aphid ( Melanaphis sacchari ) and head smut fungi ( Sporisorium reilianum ) infesting sorghum cause delayed growth and development, and reduce yield and quality. This study use bioinformatics and molecular biological approaches to profile the gene expression pattern during sorghum development and under pest infestation, and analyzed the natural allelic DNA variation of sorghum MYC gene family. The findings provide insights for potential application in breeding the stress resistant and high productivity sorghum varieties. The results indicated that there are 28 MYC genes identified in sorghum genome, distributed on 10 chromosomes. The bHLH_MYC_N and HLH domains are the conserved domains of the MYC gene in sorghum. Gene expression analysis showed that SbbHLH35 . 7g exhibited high expression levels in leaves, SbAbaIn showed strong expression in early grains, and SbMYC2 . 1g showed high expression levels in mature pollen. In anti-aphid strains at the 5-leaf stage, SbAbaIn , SbLHW . 4g and SbLHW . 2g were significantly induced in leaves, while SbbHLH35 . 7g displayed the highest expression level in panicle tissue, which was significantly induced by the infection of head smut. Promoter cis -element analysis identified methyl jasmonate (MJ), abscisic acid (ABA), salicylic acid (SA) and MYB-binding sites related to drought-stress inducibility. Furthermore, genomic resequencing data analysis revealed natural allelic DNA variations such as single nucleotide polymorphism (SNP) and insertion-deletion (INDEL) for the key SbMYCs . Protein interaction network analysis using STRING indicated that SbAbaIn interacts with TIFYdomain protein, and SbbHLH35.7g interacts with MDR and imporin. SbMYCs exhibited temporal and spatial expression patterns and played vital roles during the sorghum development. Infestation by sugarcane aphids and head smut fungi induced the expression of SbAbaIn and SbbHLH35 . 7g , respectively. SbAbaIn modulated the jasmonic acid (JA) pathway to regulate the expression of defensive genes, conferring resistance to insects. On the other hand, SbbHLH35 . 7g participated in detoxification reactions to defend against pathogens.

Nitrate is the main form of inorganic nitrogen that crop absorbs, and nitrate transporter 2 (NRT2) is a high affinity transporter using nitrate as a specific substrate. When the available nitrate is limited, the high affinity transport systems are activated and play an important role in the process of nitrate absorption and transport. Most NRT2 cannot transport nitrates alone and require the assistance of a helper protein belonging to nitrate assimilation related family (NAR2) to complete the absorption or transport of nitrates. Crop nitrogen utilization efficiency is affected by environmental conditions, and there are differences between varieties, so it is of great significance to develop varieties with high nitrogen utilization efficiency. Sorghum bicolor has high stress tolerance and is more efficient in soil nitrogen uptake and utilization. The S . bicolor genome database was scanned to systematically analyze the gene structure, chromosomal localization, physicochemical properties, secondary structure and transmembrane domain, signal peptide and subcellular localization, promoter region cis -acting elements, phylogenetic evolution, single nucleotide polymorphism (SNP) recognition and annotation, and selection pressure of the gene family members. Through bioinformatics analysis, 5 NRT2 gene members (designated as SbNRT2 - 1a , SbNRT2 - 1b , SbNRT2 - 2, SbNRT2 - 3, and SbNRT2 - 4 ) and 2 NAR2 gene members (designated as SbNRT3 - 1 and SbNRT3 - 2 ) were identified, the number of which was less than that of foxtail millet. SbNRT2 / 3 were distributed on 3 chromosomes, and could be divided into four subfamilies. The genetic structure of the same subfamilies was highly similar. The average value of SbNRT2/3 hydrophilicity was positive, indicating that they were all hydrophobic proteins, whereas α-helix and random coil accounted for more than 70% of the total secondary structure. Subcellular localization occurred on plasma membrane, where SbNRT2 proteins did not contain signal peptides, but SbNRT3 proteins contained signal peptides. Further analysis revealed that the number of transmembrane domains of the SbNRT2s family members was greater than 10, while that of the SbNRT3s were 2. There was a close collinearity between NRT2 / 3s of S . bicolor and Zea mays . Protein domains analysis showed the presence of MFS_1 and NAR2 protein domains, which supported executing high affinity nitrate transport. Phylogenetic tree analysis showed that SbNRT2 / 3 were more closely related to those of Z . mays and Setaria italic . Analysis of gene promoter cis -acting elements indicated that the promoter region of SbNRT2 / 3 had several plant hormones and stress response elements, which might respond to growth and environmental cues. Gene expression heat map showed that SbNRT2 - 3 and SbNRT3 - 1 were induced by nitrate in the root and stem, respectively, and SbNRT2 - 4 and SbNRT2 - 3 were induced by low nitrogen in the root and stem. Non-synonymous SNP variants were found in SbNRT2 - 4 and SbNRT2 - 1a . Selection pressure analysis showed that the SbNRT2 / 3 were subject to purification and selection during evolution. The expression of SbNRT2 / 3 gene and the effect of aphid infection were consistent with the expression analysis results of genes in different tissues, and SbNRT2 - 1b and SbNRT3 - 1 were significantly expressed in the roots of aphid lines 5-27sug, and the expression levels of SbNRT2 - 3 , SbNRT2 - 4 and SbNRT3 - 2 were significantly reduced in sorghum aphid infested leaves. Overall, genome-wide identification, expression and DNA variation analysis of NRT2/3 gene family of Sorghum bicolor provided a basis for elucidating the high efficiency of sorghum in nitrogen utilization.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a lasting threat to public health. To minimize the viral spread, it is essential to develop more reliable approaches for early diagnosis of the infection and immediate suppression of the viral replication. Herein, through computational prediction of SARS-CoV-2 genome and screening analysis of specimens from covid-19 patients, we predicted 15 precursors for SARS-CoV-2-encoded miRNAs (CvmiRNAs) containing 20 mature CvmiRNAs, in which CvmiR-2 was successfully detected by quantitative analysis in both serum and nasal swab samples of patients. CvmiR-2 showed high specificity in distinguishing covid-19 patients from normal controls, and high conservation between SARS-CoV-2 and its mutants. A positive correlation was observed between the CvmiR-2 expression level and the severity of patients. The biogenesis and expression of CvmiR-2 were validated in the pre-CvmiR-2-transfected A549 cells, showing a dose-dependent pattern. The sequence of CvmiR-2 was validated by sequencing analysis of human cells infected by either SARS-CoV-2 or pre-CvmiR-2. Target gene prediction analysis suggested CvmiR-2 may be involved in the regulation of the immune response, muscle pain and/or neurological disorders in covid-19 patients. In conclusion, the current study identified a novel v-miRNA encoded by SARS-CoV-2 upon infection of human cells, which holds the potential to serve as a diagnostic biomarker or a therapeutic target in clinic., 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 © 2023 Zhao, Lü, Zhao, Guo, Wang, Yu, Hao, Zhu and Yu.)