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.

Exploit SSR in Cryptromeria fortunei and application in genetic diversity analysis

Cryptromeria fortunei is one of the main forest plantation species in the subtropical high altitude areas in China. In this paper we collected 49 C. fortunei germplasm resources and provides a study of the utility of freely available C. japonica EST resources for the development of markers necessary for genetic diversity analyses of C. fortunei. By screening 24,299 EST sequences from C. japonica with SSR Finder, we identified 2384 ESTs car- rying 2783 SSR motifs. We successfully obtained 364 （15 %） primers from 2419 putative SSR loci. Of the 80 candidate SSR markers tested, 70 （87.5 %） yielded stable and clear PCR products. With those primers, the genetic diversity of 49 C. fortunei we collected was studied. The results showed that 18 primers yield polymorphism within these accessions. These 18 primers generated 48 scorable SSR loci and the average number of polymorphic SSR loci per primer was 2.7. The PIC value varied from 0.375 to 0.8101, with the average of 0.4780. The Shannon index is 0.5718, and the value of the observed number of alleles and effective number of alleles are 1.9167 and 1.7289, respectively. The genetic coefficient of these 49 accessions varied from 0.28 to 0.87. According to the genetic dis- tances, a cluster tree was constructed. At genetic coefficient of 0.60, these 49 accessions can group into 3： group I contains only FJ-laizhou accession, and group II contains 2accessions from FJ-layang, and the other one group con- tains mixed accessions. At genetic coefficient of 0.68, the former group II was constructed into 7 subgroups, with the first 3 subgroups contain 16 accessions in which 11 （69 %） are from Fujian province, and the later 4 subgroup contain 31 accessions in which 20 （65 %） were from Zhejiang province.

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.

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.