Genome-wide identification,characterization, and expression analysis of nucleotide-binding leucine-rich repeats gene family under environmental stresses in tea (Camellia sinensis)

Plants often use nucleotide-binding leucine-rich repeats (NLRs) to recognize specific virulence proteins and activate the hypersensitive response thereby defending against invaders. However, data on NLRs and the resistance mechanism of NLR protein mediation in tea plant are extremely limited. In this study, 400 and 303 CsNLRs were identified from the genomes of C. sinensis var. sinensis (CSS) and C. sinensis var. assamica (CSA), respectively. Phylogenetic analysis revealed that the numbers in CNL groups are predominant in both CSS and CSA. RNA-Seq revealed that the expression of CsNLRs is induced by Colletotrichum fructicola, cold, drought, salt stress and exogenous methyl jasmonate. The 21 CsCNLs that are highly expressed in tea plant under biotic and abiotic stresses as well as during bud dormancy and in different tissues are identified. Gene structure analysis revealed several cis-regulatory elements associated with phytohormones and light responsiveness in the promoter regions of these 21 CsCNLs.     

Validation of reliability for reference genes under various abiotic stresses in tea plant

Reference genes are frequently used as a normalization standard to obtain reliable data during quantitative real-time polymerase chain reaction (qRT-PCR). However, recent studies showed that most traditional reference genes were not stable under different treatments or environmental stresses, which may lead to misinterpret expression of the target genes. In this study, 7 candidate reference genes in tea plant (Camellia sinensis (L.) Kuntze cv. Yingshuang) were selected and their expression stability under different abiotic stresses was analyzed using geNorm, NormFinder, and BestKeeper methods. Our results suggest that TUA1 (alpha-1 tubulin) has the most stable expression under damage stresses according to 3 methods of analysis. For drought stresses, 18S rRNA, and GAPDH (glyceraldehyde-3-phosphate dehydrogenase) were the most stable genes. For cold, Al, and NaCl stresses, GAPDH and TUA1 may be the alternative options. Our results may provide an insight for identification of the optimal reference genes for tea plants under various treatments.