Whole-genome re-sequencing, diversity analysis, and stress-resistance analysis of 77 grape rootstock genotypes.

Introduction: Grape rootstocks play critical role in the development of the grape industry over the globe for their higher adaptability to various environments, and the evaluation of their genetic diversity among grape genotypes is necessary to the conservation and utility of genotypes. Methods: To analyze the genetic diversity of grape rootstocks for a better understanding multiple resistance traits, whole-genome re-sequencing of 77 common grape rootstock germplasms was conducted in the present study. Results: About 645 billion genome sequencing data were generated from the 77 grape rootstocks at an average depth of ~15.5×, based on which the phylogenic clusters were generated and the domestication of grapevine rootstocks was explored. The results indicated that the 77 rootstocks originated from five ancestral components. Through phylogenetic, principal components, and identity-by-descent (IBD) analyses, these 77 grape rootstocks were assembled into ten groups. It is noticed that the wild resources of V. amurensis and V. davidii, originating from China and being generally considered to have stronger resistance against biotic and abiotic stresses, were sub-divided from the other populations. Further analysis indicated that a high level of linkage disequilibrium was found among the 77 rootstock genotypes, and a total of 2,805,889 single nucleotide polymorphisms (SNPs) were excavated, GWAS analysis among the grape rootstocks located 631, 13, 9, 2, 810, and 44 SNP loci that were responsible to resistances to phylloxera, root-knot nematodes, salt, drought, cold and waterlogging traits. Discussion: This study generated a significant amount of genomic data from grape rootstocks, thus providing a theoretical basis for further research on the resistance mechanism of grape rootstocks and the breeding of resistant varieties. These findings also reveal that China originated V. amurensis and V. davidii could broaden the genetic background of grapevine rootstocks and be important germplasm used in breeding high stress-resistant grapevine rootstocks.

Transcriptomic Analysis Elaborates the Resistance Mechanism of Grapevine Rootstocks against Salt Stress.

Grapes are subject to a wide range of climatic conditions during their life cycle, but the use of rootstocks can effectively ameliorate the effects of abiotic stress. However, the tolerance mechanism of different grape rootstock varieties varies under various stresses, and systematic research on this aspect is limited. On the basis of previous research, transcriptome sequencing was performed on three tolerant grape rootstock varieties (3309C, 520A, 1103P) and three intolerant grape rootstock varieties (5BB, 101-14, Beta). In total, 56,478,468 clean reads were obtained. One hundred and ten genes only existed in all combinations during P1 with a downregulated trend, and 178 genes existed only in P1 of tolerant grape rootstock varieties. Salt treatment firstly affected the photosynthesis of leaves, and tolerant varieties weakened or even eliminated this effect through their own mechanisms in the later stage. Tolerant varieties mobilized a large number of MFs during the P2 stage, such as hydrolase activity, carboxypeptidase activity, and dioxygenase activity. Carbon metabolism was significantly enriched in P1, while circadian rhythm and flavonoid biosynthesis were only enriched in tolerant varieties. In the intolerant varieties, photosynthesis-related pathways were always the most significantly enriched. There were large differences in the gene expression of the main signal pathways related to salt stress in different varieties. Salt stress affected the expression of genes related to plant abiotic stress, biotic stress, transcription factors, hormones, and secondary metabolism. Tolerant varieties mobilized more bHLH, WRKY, and MYB transcription factors to respond to salt stress than intolerant varieties. In the tolerant rootstocks, SOS was co-expressed. Among these, SOS1 and SOS2 were upregulated, and the SOS3 and SOS5 components were downregulated. The genes of heat shock proteins and the phenylalanine pathway were upregulated in the tolerant varieties. These findings outline a tolerance mechanism model for rootstocks for coping with osmotic stress, providing important information for improving the resistance of grapes under global climate change.

Cellular autofluorescence and browning in trichomes of Chinese cabbage (Brassica campestris) in response to mechanical stimulation and senescence.

There is limited information concerning the formation of dot-like browning appearing at the base of trichomes on mature leaves on the Chinese cabbage (Brassica campestris L. ssp. pekinensis). This study confirmed for the first time that enhanced autofluorescence can be induced in the base of trichomes when pressure stimuli is applied to trichomes; the enhanced autofluorescence gradually moves to the top of trichomes and the neighbouring mesophyll tissue within 15min. The excitation of autofluorescence in trichomes was found to be more effective in mature leaves compared to newly emergent leaves. Increased polyphenol oxidase (PPO) activities and reactive oxygen species (ROS) accumulation were also detected in the basal region of trichomes that were subjected to mechanical stimuli. Enhanced fluorescence was observed at the top of the trichomes in senescencing leaves. A browning in the base of the trichomes during leaf senescence was observed. In contrast, no browning occurred at the base of the trichomes in leaves that were subject to pressure stimuli. The blue fluorescence in the trichomes in senescent leaves arises mainly from the condensed cytoplasm. No direct evidence was able to prove that the enhanced autofluorescent substances in the trichomes during leaf senescence are the cause of the browning at the early growth stages.

[Effects of arbuscular mycorrhizal fungi on metabolism of aroma substances in tobacco]. / ä¸›æžèŒæ ¹çœŸèŒå¯¹çƒŸè‰é¦™æ°”ç›¸å ³ç‰©è´¨ä»£è°¢çš„å½±å“.

Arbuscular mycorrhizal fungi (AMF) can promote nutrient absorption and improve stress resistance of host plants. The effects of AMF on aroma substance metabolism were rarely examined. In this study, we investigated the effects of AMF (Glomus mosseae) on glands and metabolism of aroma substances in tobacco leaves. The results showed that the density of gland hair and the relative expression of the glandular-specific lipid transporter gene NtLTP1, which was necessary to induce lipid secretion, were higher in the leaves of tobacco inoculated with AMF. The content of main aroma substances in tobacco leaves, such as carotenoids, chlorogenic acid and solanesol, were increased. Moreover, AMF inoculation increased the activities of phenylalanine ammonia-lyase (PAL) and polyphenol oxidase (PPO), the key aroma substance synthesis enzymes, and caused upregulation in the relative expression of phenylalanine transaminase, polyphenoloxidase, flavonoids alcoholase and squalene synthase encoding genes. In all, the symbiosis with G. mosseae could increase the abundance and secretory activity of glandular hairs and promote the synthesis of aroma substances in tobacco leaves.

VvBAP1 Is Involved in Cold Tolerance in Vitis vinifera L.

The majority of commercial grape cultivars originate from the European grape. While these cultivars have excellent organoleptic qualities, they suffer from a relatively poor tolerance to the cold experienced during winter, resulting in significant damage to grapevines. Thus, low temperature is one of the bottlenecks that restrict the further growth of the grape industry. Research on the mechanism of cold tolerance in grapes is therefore very important. BON association protein 1 (BAP1) is a recently discovered phospholipid-binding protein. In Arabidopsis, the expression of AtBAP1 can be regulated via low temperature; however, the function of BAP1 in the grapevine has not been reported. The VvBAP1 gene was cloned in our previous studies in grapes, and bioinformatics analysis showed that it harbors the conservative calcium-dependent C2 protein domain. However, little is known about its function and underlying mechanism. In this study, cold treatment was applied to the cold-resistant grape varieties 'F-242' and 'Zuoyouhong' as well as to the cold-sensitive grape varieties 'Cabernet Sauvignon' and 'Chardonnay.' The expression level of VvBAP1 in the cold-resistant varieties was significantly higher than in the cold-sensitive varieties, indicating that VvBAP1 could be associated with the cold response processes in the grapevine. Using the cold-resistant grape variety 'F-242' as material, with the 4°C and CaCl2 treatment, the relative expression of VvBAP1 was determined via qRT-PCR. Both low temperature and low-temperature signal Ca2+ induced VvBAP1 expression. In addition, the VvBAP1 gene was cloned and transferred into Arabidopsis to generate VvBAP1 overexpressing plants. Biochemical assays and gene expression analyses were conducted on plants subjected to low temperature treatments (4 and -8°C). The obtained results showed that the activities of superoxide dismutase and peroxidase in these transgenic plants were higher than those in wild type (WT) plants, and that cell membrane permeability and malondialdehyde content were both lower compared to WT plants. Furthermore, the content of soluble sugars and the expression levels of sugar-metabolizing related genes, such as BAM4-7, SS4, and G6PD5, were significantly higher than those of WT plants. Furthermore, the expression of low temperature response signal genes, including CBF1, CBF3, COR15a, COR6.6, COR27, and KIN1, were also enhanced. In summary, these results showed that VvBAP1 could strengthen the cold resistance in the grapevine through adjusting and controlling the sugar content and activating antioxidant enzyme activity.

Characterization of miRNAs responsive to exogenous ethylene in grapevine berries at whole genome level.

MicroRNAs (miRNAs) are critical regulators of various biological and metabolic processes of plants. Numerous miRNAs and their functions have been identified and analyzed in many plants. However, till now, the involvement of miRNAs in the response of grapevine berries to ethylene has not been reported yet. Here, Solexa technology was employed to deeply sequence small RNA libraries constructed from grapevine berries treated with and without ethylene. A total of 124 known and 78 novel miRNAs were identified. Among these miRNAs, 162 miRNAs were clearly responsive to ethylene, with 55 downregulated, 59 upregulated, and 14 unchanged miRNAs detected only in the control. The other 35 miRNAs responsive to ethylene were induced by ethylene and detected only in the ethylene-treated grapevine materials. Expression analysis of 27 conserved and 26 novel miRNAs revealed that 13 conserved and 18 novel ones were regulated by ethylene during the whole development of grapevine berries. High-throughput sequencing and qRT-PCR assays revealed consistent results on the expression results of ethylene-responsive miRNAs. Moreover, 90 target genes for 34 novel miRNAs were predicted, most of which were involved in responses to various stresses, especially like exogenous ethylene treatment. The identified miRNAs may be mainly involved in grapevine berry development and response to various environmental conditions.

[The role of nitric oxide in ethylene-induced stomatal closure in Vicia faba L].

The effects of nitric oxide (NO) and ethylene on Vicia faba L. stomatal movement were studied. The results showed that NO donor SNP (sodium nitroprusside) 10 micromol/L and ethylene 0.04% could induce stomatal closure distinctly and they could promote stomatal closure when treated together. When treated with AVG (an inhibitor of ethylene synthesis), c-PTIO (a specific scavenger of NO) and NaN(3) (an inhibitor of NR), the effects of NO- and ethylene-induced stomatal closure were inhibited but the inhibitor of nitric oxide synthase (NOS) had little effect. We presumed that there was coordinative effect between NO and ethylene in regulation of stomatal closure; ethylene could induce stomatal closure by regulating the production of nitrate reductase (NR)-dependent NO.

[Involvement of nitric oxide in regulation of salt stress-induced ABA accumulation in maize seedling].

Possible regulation of salt stress-induced ABA accumulation by nitric oxide (NO) in maize seedling was investigated. Both NO and ABA contents of maize leaves and root tips were increased in response to salt stress (Fig.1,2). Similar to the effects of salt stress, ABA contents of maize leaves and root tips were increased after the treatment of maize leaves with sodium nitroprusside (SNP, a nitric oxide donor) alone (Fig.3). Compared to the salt stress-induced ABA accumulation, this SNP-induced ABA increase was much faster, suggesting that NO may be an intermediate signal from salt stress to ABA accumulation. When NO production inhibitors L-NAME and NaN(3) treatments were applied, salt stress-induced ABA accumulation was lowered (Fig.4). Treatment with NO scavenger cPTIO also inhibited the salt stress-induced ABA increase (Fig.5). From these results it is deduced that NO is involved in regulation of ABA accumulation under salt stress.