Gene expression profiling reveals the effects of light on adventitious root formation in lotus seedlings (Nelumbo nucifera Gaertn.)

Abstract

Background Lotus is an aquatic horticultural crop that is widely cultivated in most regions of China and is used as an important off-season vegetable. The principal root of lotus is degenerated, and adventitious roots (ARs) are irreplaceable for plant growth. We found that no ARs formed under darkness and that exposure to high-intensity light significantly promoted the development of root primordia. Four differential expression libraries based on three light intensities were constructed to monitor metabolic changes, especially in indole-3-acetic acid (IAA) and sugar metabolism. Results AR formation was significantly affected by light, and high light intensity accelerated AR development. Metabolic changes during AR formation under different light intensities were evaluated using gene expression profiling by high-throughput tag-sequencing. More than 2.2 × 104 genes were obtained in each library; the expression level of most genes was between 0.01 and 100 (FPKF value). Libraries constructed from plants grown under darkness (D/CK), under 5000 lx (E/CK), and under 20,000 lx (F/CK) contained 1739, 1683, and 1462 upregulated genes and 1533, 995, and 834 downregulated genes, respectively, when compared to those in the initial state (CK). Additionally, we found that 1454 and 478 genes had altered expression in a comparison of libraries D/CK and F/CK. Gene transcription between libraries D/F ranged from a 5-fold decrease to a 5-fold increase. Twenty differentially expressed genes (DEGs) were involved in the signal transduction pathway, 28 DEGs were related to the IAA response, and 35 DEGs were involved in sugar metabolism. We observed that the IAA content was enhanced after seed germination, even in darkness; this was responsible for AR formation. We also observed that sucrose could eliminate the negative effect of 150 μMol IAA during AR development. Conclusions AR formation was regulated by IAA, even in the dark, where induction and developmental processes could also be completed. In addition, 36 genes displayed altered expression in carbohydrate metabolism and ucrose metabolism was involved in AR development (expressed stage) according to gene expression and content change characteristics.

The lotus NnWOX1-1、NnWOX4-3 and NnWOX5-1 promoted root formation and enhanced stresses tolerance in transgenic Arabidopsis thaliana

“…The same phenomenon is also found in transgenic WOX 11 and WOX 12 plants, transgenic plantgs have more root number than that of none transgenic plant [46,47]. In previous study, we observed that the expression of some NnWOXs were induced by IAA and sucrose through screening transcripts in the whole genome of louts [20][21]. In this study three genes including NnWOX1-1, NnWOX4-3 and NnWOX5-1 were cloned with RT-PCR method (additional Fig.…”

“…β-Actin of lotus was used as an internal standard, and the forward: 5'-AACCTCCTCCTCATCGTACT-3', and reverse: 5'-GACAGCATCAG CCATGTTCA-3'. the mRNA level was calculated with 2 −△△Ct method [21]. A total of 25 µl reaction mixture containing 12.5 µl SYBR Premix Ex Taq II (Tli RNaseH Plus) (2×), 1 µl of each primers (forward and reverse), 3 µl cDNAs of NnWOX1-1, NnWOX4-3 and NnWOX5-1, and 8.5 µl dH 2 O was prepared.…”

“…About 2-3 µg RNA of each material were prepared for library construction. The detail process for each library construction was referred by Cheng et al (2020a) [21], and sequencing by Nanjing Jisi Huiyuan Biotechnology Co., Ltd with a special construct.…”

“…The supernatant at lower layer was selected, and dried with nitrogen. The dry powder was dissolved by 100 ml ltered methanol, and 50 of above solution was transferred into liquid chromatography (Sigma, Germany) for IAA content determination [21]. The process of ABA content identi cation was refered to the method reported by Zdunek and Lips (2001) [43].…”

“…Low concentration of IAA promotes the cell differentiation, and high concentration of exogenous IAA results in adverse effect [20]. Light and photosynthetic products (sucrose) is also found to have a great in uence on the formation of ARs [21,22]. It is reported that lignin is involved in lotus ARs formation, which is also regulated by sucrose, suggesting that the development of ARs also has a complex regulatory network in lotus [23].…”