- Genome-wide analysis of long non-coding RNAs affecting roots development at an early stage in the rice response to. - Background: Long non-coding RNAs (lncRNAs) have been found to play a vital role in several gene regulatory networks involved in the various biological processes in plants related to stress response. - Thus, we presented the characterization and expression of lncRNAs in rice root development at an early stage in response to Cd stress.. - Results: The lncRNA deep sequencing revealed differentially expressed lncRNAs among Cd stress and normal condition.. - In the Cd stress group, 69 lncRNAs were up-regulated and 75 lncRNAs were down-regulated. - The pathway analyses found that secondary metabolites, such as phenylpropanoids and phenylalanine, and photosynthesis pathway-related genes were significantly altered by Cd stress.. - It also expands our knowledge about lncRNA biological function and contributes to the annotation of the rice genome.. - LncRNAs are also involved in the regulation of targeted genes via epi- genetic, transcriptional and post-transcriptional methods. - Although previous studies have provided useful information on the mechanisms for rice lncRNA in Cd stress response, the regulatory mech- anisms involved are mostly unknown. - To improve our understanding of the likely functional roles of lncRNAs in rice Cd stress response, further studies are necessary to understand the functional genetics of lncRNAs in detail, and to determine which specific lncRNAs target selective sites for interaction in the rice genome.. - In the current study, we used a deep RNA sequencing strategy to clarify the lncRNAs profiled associated with Cd stress using the Cd response rice genotype which could provide more insights into the regulatory role of more lncRNAs in the rice Cd stress response. - The results ob- tained in our study provided a valuable resource to study the lncRNAs involved in Cd stress response and will in- crease the knowledge for a better understanding of the biological processes of rice stress response.. - DX142 is a pure line and shows the highest sensitivity to Cd stress.. - NaClO solution for 30 min, rinsed five times with dis- tilled water and maintained for 2 days at 25–30 °C in the dark, thereby inducing germination. - RNA isolation, quantification and library preparation for lncRNA sequencing, clustering and sequencing and quality control were analysed as described by Ren et al.. - Fi- nally, the lncRNAs developed were referred to as Cd stress (SY_R) and control (CK_R)-related lncRNAs.. - Identification of differentially expressed lncRNAs. - Morphology under cd stress and control conditions First, 82 different rice genotypes were used in this study to check their Cd stress sensitivity for acquiring add- itional insights into the rice transcriptomic response to environmental Cd stress. - The average root length of DX142 under Cd stress and control conditions is significantly different. - The average root length of DX142 under the control condition was significantly higher than under the Cd stress condition from the second day (Fig. - Identification of lncRNAs. - After screening using harsh criteria and two analytic tools, a total of 2580 lncRNAs from Cd stress and control conditions in rice were identified and sub- jected for further analysis (Fig. - Genome-wide analysis of lncRNA expression under Cd stress and control conditions was performed to profile differentially expressed lncRNAs associated with Cd stress.. - The cis role of differentially expressed lncRNAs in target genes. - We examined protein- coding genes 10 and 100 kb upstream and downstream of the lncRNAs, respectively. - one of them is the cysteine and me- thionine metabolism pathway, which was significant in the Cd stress condition compared with the normal con- dition with an enrichment (q <. - The trans role of lncRNAs in target genes. - The pathway analyses revealed 118 different pathways corresponding to the target genes, and four of them are biosynthesis of secondary metabolites, phenylpropanoid biosynthesis, phenylalanine metabolism and photosyn- thesis signalling pathways in the Cd stress condition. - Table 1 Statistics of the read alignments in the RNA-Seq study. - We randomly se- lected 10 genes in the lncRNA-seq data. - The study of differentially expressed lncRNAs control- ling gene expression in the rice stress response at the whole transcriptome level especially in Cd stress, is re- ported to be limited. - Thus, our data systemically predict the lncRNAs at the whole transcriptome level and showed that which specific lncRNAs seek out selective sites in the genome for interaction in rice Cd stress. - Here, 143 lncRNA genes were detected that showed dif- ferential expression in the Cd stress response in the root, and 83.9 and 100% of them were identified that contained at least one differentially expressed mRNA in cis and trans, respectively, which suggests that these lncRNAs may play an important role in the rice Cd stress response. - Further- more, lncRNAs showed a higher ratio than those of the other two subtypes of all the differentially expressed. - 2 Screening of the candidate lncRNAs. - lncRNAs, indicating that lncRNAs may be the main form of lncRNAs in the Cd stress response in rice. - major form of regulation in the Cd stress process for rice.. - In addition, we compared the lncRNA data with the study of Fei et al. - Therefore, lncRNAs should be given more attention in heavy metal stress response studies in the future.. - KEGG analysis showed a significant change in the cysteine and methionine metab- olism pathway in the Cd stress compared to normal condi- tion. - 4 Scatter plot of KEGG pathway enrichment statistics for differentially expressed target genes in cis in rice roots. - Table 2 Significant pathways and proportions after KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis of differentially expressed target genes in cis in the root. - chelate Cd to procedure non-toxic complexes which are then sequestered into the vacuole to avoid high levels of free cytotoxic Cd in the cytosol in response to Cd stress [53]. - In this study, we observed that OS03G0196600, which is involved in the cysteine and methionine metabolism pathways, was clearly up-regulated (Additional file 9: Table S5) and might contribute to the production of cysteine-rich (Cys-rich) peptides. - Previous studies showed that expos- ure to Cd stress will lead to impairment of the photosyn- thetic function in many plant species. - Furthermore, ABA was found to be involved in the regulation of antioxida- tive defence systems and Cd-induced oxidative stress in mung bean seedlings [58]. - In the current study, the trans role of lncRNAs, including XLOC_058523, XLOC_104363 and XLOC_059778, targeted phenylpropanoids and the phenyl- alanine related-gene OS11G0552000, which indicated that lncRNAs may regulate the genes of the secondary metabo- lites in far distance and then activate the various trans- porters to successively guide removal of excessive Cd from the cell. - (Kyoto Encyclopedia of Genes and Genomes) analysis of differentially expressed target genes in trans in the root. - 5 Gene Ontology (GO) enrichment analysis for differentially expressed target genes in trans in rice roots. - The y-axis and x-axis indicate the number of genes in a category and the names of the clusters, respectively. - A similar result can be seen in the study of Zhai et al. - These results suggest that the photosyn- thesis pathway-related genes are involved in the Cd stress response in rice and may also be regulated by lncRNAs in trans. - In addition, after pathway analyses of the target genes of these differ- entially expressed lncRNAs, cysteine and methionine me- tabolism pathway, carotenoid biosynthesis, ABA signalling pathway (in cis), and secondary metabolites and photosyn- thesis (in trans) were enriched, which indicated that lncRNAs may play an important role in these pathways in response to Cd stress. - Therefore, further studies are ne- cessary needed to fully understand these lncRNAs to ef- fectively control rice Cd pollution in the future.. - Differentially expressed lncRNAs in response to Cd stress in root libraries. - native mean that the gene is up-regulated and down-regulated in the CK sample, respectively. - Pathways and proportions after KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis of differentially expressed target genes in cis in the root. - Pathways and proportions after KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis of differentially expressed target genes in trans in the root. - We made a random selection of 10 genes in the genes ’ data of lncRNA-seq. - Comparison of the log 2 (FC) of 10 selected transcripts using RNA-Seq and qRT-PCR. - The differentially expressed mRNAs by lncRNA-seq. - lncRNAs: Long non-coding RNAs. - SY_R: The rice root in Cd stress Fig. - The y-axis and x-axis indicate Log 2 (Fold Change) and the name of the genes, respectively. - XLOC_033045 represent one of the whole lncRNAs. - The funding body was not involved in the design of the study, analysis or interpretation of data or writing the manuscript.. - The sequence data generated during the current study are available in the NCBI Sequence Read Archive under the accession number of SRP099996 (https://www.ncbi.nlm.nih.gov/sra/SRP099996).. - The rice line DX142 (including the seeds) obtained from the native cultival line and all plants were grown in the test fields of Jiangxi Agriculture University.. - 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