Response of Chinese fir seedlings to low phosphorus stress and analysis of gene expression differences

Chinese fir(Cunninghamia lanceolata) is an excellent fast-growing timber species occurring in southern China and has significant value in the forestry industry. In order to enhance the phosphorus utilization efficiency in Chinese fir, four clones named X6, S3, S39 and FK were used, and low phosphorus(LP) stress experiments were performed to analyze the response of different clones to phosphorus deficiency. According to the results on seedling height, maximum root length, leaf blade aspect ratio, root ratio, malondialdehyde content, acid phosphates activity,proline content, soluble protein level, and chlorophyll a and b levels of the tested clones, compared to the control groups(CK), the phosphorus high efficiency clone X6 was screen out for transcriptome sequencing experiments. De novo RNA-seq was then used to sequence the root transcriptomes of X6 under LP stress and CK, and we then compared the gene expression differences under the two conditions. A total of 3416 SDEGs were obtained by comparing the LP and CK groups, among which 1742 were up-regulated and 1682 were down-regulated. All SDEGs obtained from the LP and CK treated samples were subjected to KEGG annotation and classification. Through classification statistical analysis using WEGO software,607 SDEGs obtained KEGG pathway annotations, which were related to 206 metabolic pathways. In Chinese fir subjected to LP stress, 53 SDEGs related with phosphorus metabolism, and phosphate uptake and transport were obtained from our transcriptome data. Based on the phosphorus metabolism pathway obtained by KEGG classification, combined with previously report on gene annotation related with phosphorus metabolism, the enzymes encoded by SDEG related with phosphorus metabolism and their expression pattern were mapped onto phosphorus metabolism pathway.

Identification and evaluation of reference genes for normalization in quantitative real-time PCR analysis in the premodel tree Betula luminifera

Betula luminifera is a commercial tree species that is emerging as a new model system for tree genomics research. A draft genomic sequence is expected to be publicly available in the near future, which means that an explosion of gene expression studies awaits. Thus, the work of selecting appropriate reference genes for q PCR normalization in different tissues or under various experimental conditions is extremely valuable. In this study, ten candidate genes were analyzed in B. luminifera subjected to different abiotic stresses and at various flowering stages.The expression stability of these genes was evaluated using three distinct algorithms implemented using ge Norm,Norm Finder and Best Keeper. The best-ranked reference genes varied across different sample sets, though RPL39,MDH and EF1 a were determined as the most stable by the three programs among all tested samples. RPL39 and EF1 a should be appropriate for normalization in N-starved roots,while the combination of RPL39 and MDH should be appropriate for N-starved stems and EF1 a and MDH should be appropriate in N-starved leaves. In PEG-treated（osmotic） roots, MDH was the most suitable, whereas EF1 a was suitable for PEG-treated stems and leaves. TUA was also stably expressed levels in PEG-treated plants. The combination of RPL39 and TUB should be appropriate for heat-stressed leaves and flowering stage. For reference gene validation, the expression levels of SOD and NFYA-3were investigated. This work will be beneficial to future studies on gene expression under different abiotic stress conditions and flowering status in B. luminifera.

Orthogroup and phylotranscriptomic analyses identify transcription factors involved in the plant cold response:Acase study of Arabidopsis BBX29

C-repeat binding factors(CBFs)are well-known transcription factors(TFs)that regulate plant cold acclimation.RNA sequencing(RNA-seq)data from diverse plant species provide opportunities to identify other TFs involved in the cold response.However,this task is challenging because gene gain and loss has led to an intertwined community of co-orthologs and in-paralogs between and within species.Using orthogroup(closely related homologs)analysis,we identified 10,549 orthogroups in five representative eudicots.A phylotranscriptomic analysis of cold-treated seedlings from eudicots identified 35 high-confidence conserved cold-responsive transcription factor orthogroups(CoCoFos).These 35 CoCoFos included the well-known cold-responsive regulators CBFs,HSFC1,ZAT6/10,and CZF1 among others.We used Arabidopsis BBX29 for experimental validation.Expression and genetic analyses showed that cold-induction of BBX29 is CBF-and abscisic acid-independent,and BBX29 is a negative regulator of cold tolerance.Integrative RNA-seq and Cleavage Under Targets and Tagmentation followed by sequencing analyses revealed that BBX29 represses a set of cold-induced TFs(ZAT12,PRR9,RVE1,MYB96,etc.).Altogether,our analysis yielded a library of eudicot CoCoFos and demonstrated that BBX29 is a negative regulator of cold tolerance in Arabidopsis.

The chromosome-scale genome of Phoebe bournei reveals contrasting fates of terpene synthase (TPS)-a and TPS-b subfamilies

Terpenoids,including aromatic volatile monoterpenoids and sesquiterpenoids,function in defense against pathogens and herbivores.Phoebe trees are remarkable for their scented wood and decay resistance.Unlike other Lauraceae species investigated to date,Phoebe species predominantly accumulate sesquiterpenoids instead of monoterpenoids.Limited genomic data restrict the elucidation of terpenoid variation and functions.Here,we present a chromosome-scale genome assembly of a Lauraceae tree,Phoebe bournei,and identify 72 full-length terpene synthase(TPS)genes.Genome-level comparison shows pervasive lineage-specific duplication and contraction of TPS subfamilies,which have contributed to the extreme terpenoid variation within Lauraceae species.Although the TPS-a and TPS-b subfamilies were both expanded via tandem duplication in P.bournei,more TPS-a copies were retained and constitutively expressed,whereas more TPS-b copies were lost.The TPS-a genes on chromosome 8 functionally diverged to synthesize eight highly accumulated sesquiterpenes in P.bournei.The essential oil of P.bournei and its main component,b-caryophyllene,exhibited antifungal activities against the three most widespread canker pathogens of trees.The TPS-a and TPS-b subfamilies have experienced contrasting fates over the evolution of P.bournei.The abundant sesquiterpenoids produced by TPS-a proteins contribute to the excellent pathogen resistance of P.bournei trees.Overall,this study sheds light on the evolution and adaptation of terpenoids in Lauraceae and provides valuable resources for boosting plant immunity against pathogens in various trees and crops.