Effects of different oleogelators on the structural properties and composition of iron walnut-oil oleogels.

In this study, we compared the quality of iron walnut oil (IWO) oleogels prepared with different oleogelators, including γ-oryzanol/ß-sitosterol (OZ-PS), γ-oryzanol/triglyceride (OZ-TC), monoglycerides (MGS), beeswax (BW), beeswax-monoglycerides (BW-MGS), and carnauba wax (CW). The physicochemical and component properties, rheological and textural parameters, macroscopic morphologies, and antioxidant capacities of the resulting oleogels were analyzed. In addition, their microscopic properties were analyzed using Fourier-transform infrared (FTIR), X-ray powder diffraction (XRD) spectroscopy, and polarized light microscopy (PLM). The results showed that the gel structures produced by different oleogelators did not change the fatty acid composition of IWO. In addition, the IWO oleogel prepared with OZ-PS had a more stable network structure, excellent hardness at 4℃ (1116.51 g), better antioxidant capacity (766.50 µmol TE/kg) and higher total phenolic content (14.98 mg/kg) than any other experimental IWO oleogels. Moreover, comprehensive ranking by principal component analysis of numerous characteristics showed that the OZ-PS oleogel (2.533) ranked first among the six oleogels studied. Therefore, the IWO oleogel prepared with OZ-PS is a promising product, and our results provide guidance for the preparation of IWO oleogels, such as to increase their applications in the food industry.

Genetic imprints of grafting in wild iron walnut populations in southwestern China.

BACKGROUND: Anthropogenic activities are causing unprecedented loss of genetic diversity in many species. However, the effects on genetic diversity from large-scale grafting onto wild plants of crop species are largely undetermined. Iron walnut (Juglans sigillata Dode) is a deciduous nut tree crop endemic to southwestern China with a long history of cultivation. Due to the rapid expansion of the walnut industry, many natural populations are now being replaced by cultivars grafted onto wild rootstocks. However, little is known about the potential genetic consequences of such action on natural populations. RESULTS: We sampled the scion and the rootstock from each of 149 grafted individuals within nine wild populations of J. sigillata from Yunnan Province which is the center of walnut diversity and cultivation in China, and examined their genetic diversity and population structure using 31 microsatellite loci. Scions had lower genetic diversity than rootstocks, and this pattern was repeated in seven of the nine examined populations. Among those seven populations, AMOVA and clustering analyses showed a clear genetic separation between all rootstocks and all scions. However, the two remaining populations, both from northern Yunnan, showed genetic similarity between scions and rootstocks, possibly indicating that wild populations here are derived from feralized local cultivars. Moreover, our data indicated probable crop-to-wild gene flow between scions and rootstocks, across all populations. CONCLUSIONS: Our results indicate that large-scale grafting has been causing genetic diversity erosion and genetic structure breakdown in the wild material of J. sigillata within Yunnan. To mitigate these effects, we caution against the overuse of grafting in wild populations of iron walnut and other crop species and recommend the preservation of natural genotypes through in situ  and ex situ conservation.

Genomic characterization of the NAC transcription factors, directed at understanding their functions involved in endocarp lignification of iron walnut (Juglans sigillata Dode).

The NAC (NAM, ATAF1/2, and CUC2) transcription factors (TF), one of the largest plant-specific gene families, play important roles in the regulation of plant growth and development, stress response and disease resistance. In particular, several NAC TFs have been identified as master regulators of secondary cell wall (SCW) biosynthesis. Iron walnut (Juglans sigillata Dode), an economically important nut and oilseed tree, has been widely planted in the southwest China. The thick and high lignified shell derived endocarp tissues, however, brings troubles in processing processes of products in industry. It is indispensable to dissect the molecular mechanism of thick endocarp formation for further genetic improvement of iron walnut. In the present study, based on genome reference of iron walnut, 117 NAC genes, in total, were identified and characterized in silico, which involves only computational analysis to provide insight into gene function and regulation. We found that the amino acids encoded by these NAC genes varied from 103 to 1,264 in length, and conserved motif numbers ranged from 2 to 10. The JsiNAC genes were unevenly distributed across the genome of 16 chromosomes, and 96 of these genes were identified as segmental duplication genes. Furthermore, 117 JsiNAC genes were divided into 14 subfamilies (A-N) according to the phylogenetic tree based on NAC family members of Arabidopsis thaliana and common walnut (Juglans regia). Furthermore, tissue-specific expression pattern analysis demonstrated that a majority of NAC genes were constitutively expressed in five different tissues (bud, root, fruit, endocarp, and stem xylem), while a total of 19 genes were specifically expressed in endocarp, and most of them also showed high and specific expression levels in the middle and late stages during iron walnut endocarp development. Our result provided a new insight into the gene structure and function of JsiNACs in iron walnut, and identified key candidate JsiNAC genes involved in endocarp development, probably providing mechanistic insight into shell thickness formation across nut species.

Global Transcriptomic Analyses Provide New Insight into the Molecular Mechanisms of Endocarp Formation and Development in Iron Walnut (Juglans sigillata Dode).

Iron walnut (Juglans sigillata Dode) is a native species in southwestern China that exhibits variation in both fruit morphology and shell thickness. However, the underlying molecular processes controlling hardened endocarp development in walnut has not yet been reported. Here, we generated transcriptional profiles of iron walnut endocarp at three developmental stages using "Dapao", the most common commercial variety. Using pairwise comparisons between these three stages, a total of 8555 non-redundant differentially expressed genes (DEGs) were identified, and more than one-half of the total DEGs exhibited significant differential expression in stage I as compared with stage II or stage III, suggesting that the first stage may ultimately determine the final characteristics of the mature walnut shell. Furthermore, in the clustering analysis of the above DEGs, 3682, 2349, and 2388 genes exhibited the highest expression in stages I, II, and III, respectively. GO enrichment analysis demonstrated that the major transcriptional variation among the three developmental stages was caused by differences in cell growth, plant hormones, metabolic process, and phenylpropanoid metabolism. Namely, using the tissue-specific expression analysis and a gene co-expression network, we identified MADS-box transcription factor JsiFBP2 and bHLH transcription factor JsibHLH94 as candidate regulators of endocarp formation in the early stage, and JsiNAC56 and JsiMYB78 might play key roles in regulating the lignification process of endocarp in the late stage. This study provides useful information for further research to dissect the molecular mechanisms governing the shell formation and development of iron walnut.

Population-genomic analyses reveal bottlenecks and asymmetric introgression from Persian into iron walnut during domestication.

BACKGROUND: Persian walnut, Juglans regia, occurs naturally from Greece to western China, while its closest relative, the iron walnut, Juglans sigillata, is endemic in southwest China; both species are cultivated for their nuts and wood. Here, we infer their demographic histories and the time and direction of possible hybridization and introgression between them. RESULTS: We use whole-genome resequencing data, different population-genetic approaches (PSMC and GONE), and isolation-with-migration models (IMa3) on individuals from Europe, Iran, Kazakhstan, Pakistan, and China. IMa3 analyses indicate that the two species diverged from each other by 0.85 million years ago, with unidirectional gene flow from eastern J. regia and its ancestor into J. sigillata, including the shell-thickness gene. Within J. regia, a western group, located from Europe to Iran, and an eastern group with individuals from northern China, experienced dramatically declining population sizes about 80 generations ago (roughly 2400 to 4000 years), followed by an expansion at about 40 generations, while J. sigillata had a constant population size from about 100 to 20 generations ago, followed by a rapid decline. CONCLUSIONS: Both J. regia and J. sigillata appear to have suffered sudden population declines during their domestication, suggesting that the bottleneck scenario of plant domestication may well apply in at least some perennial crop species. Introgression from introduced J. regia appears to have played a role in the domestication of J. sigillata.

Characterization of iron walnut in different regions of China based on phytochemical composition.

Little is known about the phytochemical composition of iron walnuts. Differences in the geographical origin of iron walnuts associated with economic benefits should also be examined. In this study, the phytochemical composition (fatty acids, Vitamin E, total polyphenols and flavonoids, amino acids, and minerals) of iron walnuts in China was investigated. The results showed that there were significant differences (p < 0.05) in the phytochemical composition of iron walnut oils and flours from different regions. Positive (r > 0.5, p < 0.05) and negative (r < - 0.5, p < 0.05) correlations were found between amino acids/minerals and amino acids/oleic acid, with the highest correlation coefficient (r = 0.742, p < 0.05) between Cu and tyrosine. In addition, based on the 12 phytochemical fingerprints selected by random forest, a geographical-origin identification model for iron walnuts was established, with a corresponding correct classification rate of 96.6%. The top three phytochemical fingerprints for the geographical-origin identification of iron walnut were microelements, macroelements, and antioxidant composition, with contribution rates of 61.7%, 18.1%, and 9.9%, respectively.

Completion of the Chloroplast Genomes of Five Chinese Juglans and Their Contribution to Chloroplast Phylogeny.

Juglans L. (walnuts and butternuts) is an economically and ecologically important genus in the family Juglandaceae. All Juglans are important nut and timber trees. Juglans regia (Common walnut), J. sigillata (Iron walnut), J. cathayensis (Chinese walnut), J. hopeiensis (Ma walnut), and J. mandshurica (Manchurian walnut) are native to or naturalized in China. A strongly supported phylogeny of these five species is not available due to a lack of informative molecular markers. We compared complete chloroplast genomes and determined the phylogenetic relationships among the five Chinese Juglans using IIumina sequencing. The plastid genomes ranged from 159,714 to 160,367 bp encoding 128 functional genes, including 88 protein-coding genes and 40 tRNA genes each. A complete map of the variability across the genomes of the five Juglans species was produced that included single nucleotide variants, indels (insertions and deletions), and large structural variants, as well as differences in simple sequence repeats (SSR) and repeat sequences. Molecular phylogeny strongly supported division of the five walnut species into two previously recognized sections (Juglans/Dioscaryon and Cardiocaryon) with a 100% bootstrap (BS) value using the complete cp genomes, protein coding sequences (CDS), and the introns and spacers (IGS) data. The availability of these genomes will provide genetic information for identifying species and hybrids, taxonomy, phylogeny, and evolution in Juglans, and also provide insight into utilization of Juglans plants.