To identify an embryo specific promoter of rapeseed, 990 bp-upstream of gene BnaA09g21960D (from Brassica napus L. cv Darmor) was cloned. Bioinformation showed that the segment included several special cis-acting elements (such as ABRE, GCN4_motif and TGACG-motif) which were tissue-specific, or related to hormone and stress reactions. GUS reporter gene was used, and plant expression vector DX2181-pBnaA09g21960D was constructed. GUS gene expression was found in transgenic Arabidopsis thaliana by PCR. GUS staining assays showed that the promoter could drive GUS only in seed embryo, but not in other tissues. The results indicated the tissue-specific feature of the promoter in embryo of rapeseed.
For novel insights on Mlo gene family in Brassica plants, and for revealing spectrum of plant disease resistance to powdery mildew, Mlo genes family in Brassica species and Arabidopsis thaliana were studied. Conserved domain HMM profile was used. 123 Mlo resistant genes in Brassica rapa, B. oleracea and B. napus whole genomes were focused. Evolutionary analysis on Mlo gene family among the 4 species showed that they were clustered into 3 different subgroups. Homologous relationship showed that Mlo3 and Mlo9 genes in A. thaliana had no homologous genes in Brassica species. Among the transmembrane motifs of 123 Mlo genes, more than 7 transmembrane motifs in 71 members were found, which contained more than 12 exons. These results were consistent with typical characteristics of Mlos and might be the major functional genes among these 4 species. Conserved domains of 71 Mlos in all the species had one conserved amino acid sequence (462 aa long), which might be the main functional domain. Further comparison of Mlos in A. thaliana and Brassica species uncovered that most of Mlos in A. thaliana had experienced different expansion in Brassica species except Mlo3 and Mlo9.
In order to provide a base for evaluating whether the herbicide resistant transgene from oilseed rape could introgress into wild relative weed (wild Brassica juncea), fitness of the fourth progeny of the third backcross generation, which derived from herbicide-resistant transgenic oilseed rape and wild Brassica juncea, was studied. The fitness components of BC3mF5R, BC3pF5R and BC3mF5L, BC3pF5L (R and L denoted backcross progeny obtained with glyphosate-resistant and glufosinate-resistant transgenic oilseed rape respectively. m and p denoted backcross progeny obtained with wild B. juncea as maternal plants and paternal plants respectively) between transgenic oilseed rape and wild B. juncea in field at low (15 plants per plot) and high density (30 plants per plot) under pure and mixed stands (wild B. juncea : BC3F5=1:1) were measured. The fitness components and composite fitness of BC3F5 was fit as wild B. juncea in pure stands under two densities and in mixed stands at low density. Under equal proportions at high density, BC3F5R had the similar fitness components and composite fitness to wild B. juncea. However, the main fitness components and composite fitness of BC3F5L was inferior to wild B. juncea. Correlation analysis showed that there was significant correlation between main fitness components of BC3F5 and planting density. BC3F5 between glyphosate- or glufosinate-resistant transgenic oilseed rape and wild B. juncea had the strong survival ability. Moreover, the ecological risk of gene flow was higher from glyphosate-resistant transgenic oilseed rape than that from glufosinate-resistant transgenic oilseed rape. Therefore both initial hybrid from transgenic oilseed rape to wild B. juncea and backcross between wild B. juncea and F1 or subsequent generations should be prevented to avoid producing higher fitness backcross generation.
To explore the structure of Brassica napus MAGIC (multi-parent advanced generation inter-cross) population, genetic diversity and genetic distance of the B. napus MAGIC (composed of 554 lines) were initially illustrated by 60 pairs of SSR (simple sequence repeat) primers. Among these 60 pairs, a total of 261 alleles were detected, with 4.35 alleles per pair, and the average PIC (polymorphism information content) value of 0.48. Population structure analysis using STRUCTURE 2.3.4 showed that this B. napus MAGIC population was divided into 2 subpopulations, with 303 lines in subpopulation 1 and 234 lines in subpopulation 2, and the remaining 17 lines had no specific subpopulation attribution, belonging to mixed subgroups. The results suggested that the MAGIC population had high genetic diversity.
Genetic diversities of 106 peanut varieties released from 1982 to 2016 in Henan Province were analyzed by pedigree and the coefficient of parentage (COP). The results indicated that the majority of the pedigrees of released varieties could be traced back to the same ancestor Fuhuasheng, and about 92% of the cross combinations which generated the released varieties shared parental relationship. The COP values ranged from 0 to 0.773 with the average of 0.149. Cluster analysis suggested that 3 of 106 peanut varieties were clustered into 3 groups because they shared no parental relationship with any other varieties, 13 varieties had distant parental relationship with other varieties. The remaining 90 varieties were grouped into 10 clusters, each of which consisted of certain cultivar and its derived lines. Above results indicated that the genetic background of peanut varieties released in Henan Province was quite narrow and the genetic diversity was relatively poor. Therefore, utilizing novel germplasm resources, innovating parental combinations, and enhancing the genetic diversity should be the focus of peanut breeding in future.
A total of 128 lines derived from a cross between a susceptible soybean cultivar Hefeng 25 and a disease resistant soybean cultivar Maple Arrow, were analyzed through isolated leaf stalk inoculation method and living stem inoculation method. A total of 7 QTLs were detected through combining 2 sets of phenotypic identification data with the interval mapping method. QTL for the soybean with pests or disease-resistance was collected through meta-analysis, the consistent QTL map for the soybean with pests or disease-resistance was constructed, and 2 consistent QTL were obtained. The more reduced confidence interval was obtained and 21 candidate genes were discovered, 3 of which were associated with disease resistance. The consistent results were similar to that of this study in B1 linkage group, which provided a research basis for fine positioning and gene cloning.
In this study, the soybean root specific promoter, seed specific promoter and seed coat specific promoter were cloned by homologous sequence method respectively, and their size were 2 500 bp, 1 832 bp, 1 268 bp, and they had different expression elements such as CANNTG-motifs, GATA-box, ACGT. Three plant expression vectors of these tissue specific promoters were constructed and transferred into Nicotiana tabacum NC89 by Agrobacterium-mediated method to prove GUS activity of different promoters and analyze the expression efficiency. Soybean root, seed coat and seed specific promoter nucleotide sequence were isolated by PCR method. Then 11 root specific promoter transgenic plants, 4 seed coat specific promoter transgenic plants, 8 seed specific promoter transgenic plants, and 7 35S-promoter-transgenic plants had been obtained. GUS activity assays indicated that GUS gene expression level in root, seed and seed coat were higher than that of leaf, shoot and flower. Further quantitative real time PCR results showed that the expression levels of GUS gene of root specific promoter in root was higher than in stem, leaf, seed, seed coat and flower; the expression levels of GUS gene of seed coat specific promoter in seed coat was higher than in root, stem, leaf, seed and flower; the expression levels of GUS gene of seed specific promoter in seed was higher than in root, stem, leaf, seed coat and flower, but compared to 35S promoter, the expression levels of GUS gene of these three tissue specific promoters were lower than 35S promoter in each tissues.
Inositol oxygenase is usually associated with plant cell wall synthesis and induced syncytium development. A susceptible cultivar Liaodou 15 and the resistant cultivars were Huipizhi Xiaoheidou, Harbin Xiao heidou and Xiaoli Heidou were used to study the gene expression of inositol oxygenase family gene from soybeans roots after SCN3 infection. The results showed that in both resistant cultivars the expression level of GmMIOX2 gene reached the maximum on the fifth day , but the changes of susceptible cultivar was not obvious. The expression levels of GmMIOX4 gene in Huipizhi Heidou, Harbin Xiaoheidou and Xiaoli Heidou were significantly higher than that of susceptible cultivar Liaodou 15 on fifth day, twentieth day and tenth day, indicating that GmMIOX2 and GmMIOX4 genes were related to nematode development. The complete GmMIOX2 and GmMIOX4 gene were amplified into binary expression vectors to construct pCAMBIA1303-MIOX2 and pCAMBIA1303-MIOX4. The result of subcellular localization showed that GmMIOX2 protein was located on the cytomembrane, and GmMIOX4 protein was located on cytoplasm and cytomembrane. Bioinformatic analysis of GmMIOX2 and GmMIOX4 protein domains was mainly composed of alpha helices. Phylogenetic analysis of protein sequences showed that GmMIOX2 and GmMIOX4 protein were closely related to Arabidopsis thaliana AtMIOX1 and AtMIOX4 respectively.
sesame QTL on capsule size, a genetic map was constructed based on 430 recombinant inbred lines derived from Zhongzhi 13 × ZZM2748. Capsule length, width and thickness were investigated on 2 capsule positions (middle and flanking) in 3 environments. Results showed broad ranges of length (1.54 to 4.39 cm), width (0.42 to 1.31 cm) and thickness (0.37 to 1.33 cm) respectively. Significant correlations were found between width and thickness in the capsules length and thickness of the capsules at same position. The length or width between the middle and flanking capsules also showed significant correlation. In combination with the phenotypes and genotypes of the RIL population, a total of 40 QTLs were detected with phenotype contribution rates varied from 2.00% to 23.67%. Among them, 9, 4, and 5 loci were detected for length, width and thickness of middle capsule, and 10, 8 and 4 loci were for length, width and thickness of flanking capsule. The loci qMCL13.1, qMCW9.1 and (13.77%) had the highest contribution rates (13.69% in average) to length, width and thickness of middle capsule, and qFCL11.1 (9.31%), qFCW9.1 (19.65%), qFCT9.1 (9.41%) contributed the largest variations for length, width and thickness of flanking capsule. There were 21 QTLs repeatedly observed in more than one environment and explained the phenotype variation from 2.00 to 23.67% with 9 detected across all three environments, and some of them exhibited pleiotropic effect on different traits. This study provided an insight for sesame yield related traits, and a foundation to identify capsule genes in the future.
The phenotypic variation and genetic diversity of 401 domestic and foreign flax germplasm resources were studied, by multi-statistical and SRAP marker methods. The results showed that the average variation coefficient was 17.87%, the average genetic diversity index was 2.132. There were significant differences among 28 pairs of traits. 26 pairs of SRAP primers, were used to evaluate the flax core collection, and 234 polymorphism bands were generated, with the average polymorphism information content (PIC) for each pair of primers being 0.60. The effective number of alleles (Ne), aromatic information index (I) and Nei's genetic identity (H) were 1.5847, 0.5235 and 0.3479 at the species levels, and 1.5105, 0.4644 and 0.3054 at the group levels, respectively. According to the cluster analysis, 401 flax resources could be divided into 2 categories when the genetic similarity coefficient (GS) was 0.295, and into 7 sub-types when GS was 0.375, being roughly consistent with 7 different origins. The studies showed that the agronomic traits variation was greater than that of quality traits for studied flax germplasm resources. There were great genetic differences between domestic and foreign varieties (or lines), showing a distant kinship; Geography was the main factor affecting the flax germplasm genetic diversity.
C2H2-type zinc finger protein is the key transcription factor family in plant stress resistance. A new ZAT10 gene belongs to C2H2-type zinc finger protein family was obtained from Jatropha curcas by RT-PCR, then the gene structure, functional motifs, phylogenetic relationship, and differential expression were systematically analyzed. The result showed that the full-length ORF of JcZAT10 was 753 bp, encoded a 250 amino acid polypeptide with the molecular weight of 26.7 kDa and the pI value of 8.45. JcZAT10 protein contained 2 zinc finger domains, nuclear localization signal sequence, and conserved Leu-rich motif. Its amino acid sequence shared 79.6% identity with Ricinus communis, which belonged to the same family of Euphorbiaceae. qRT-PCR analysis revealed that JcZAT10 expressed in different organs, with abundant expression in stem, but scarcely in leaves, and remarkably cold-induced expression in leaf and root, with the highest expression levels at 3 h treatment. This study laid a foundation for further studies on JcZAT10 function verification and its mechanism elucidation of cold signaling transduction in J. curcas.
Melatonin is a naturally occurring compound in plants，and this study is to reveal the effects of melatonin on soybean seedlings under salt stress. The experimental material，Tianlong 2 was planted in nutrient solution，and melatonin (200µmol/L) was applied. The traits of seedlings growth, active oxygen metabolism and osmotic adjustment substance were investigated. The results showed that salt stress inhibited the seedling growth，resulted in H2O2 and MDA accumulation，increased WSP content and enhanced the activity of SOD and POD significantly. Under salt stress condition，exogenous melatonin could alleviate the symptoms of salt toxicity and increase the biomass of soybean seedlings. The activity of antioxidant enzymes (SOD and POD) was improved significantly by exogenous melatonin，and the content of H2O2 and MDA was decreased. The results also showed that solute accumulation under stress was significantly enhanced with exogenous melatonin by increasing the content of soluble proteins. Moreover，melatonin also could promote the growth of soybean seedlings under normal conditions. This study indicated that exogenous melatonin could improve the H2O2-scavenging capacity of soybean seedlings by enhancing the activity of antioxidant enzymes and could also alleviate osmotic stress by promoting the accumulation of osmoregulatory substances. Ultimately，exogenous melatonin improved the biomass of soybean seedlings grown under salt stress，thereby effectively alleviating salt stress in soybean seedlings.
Effects of different calcium fertilizer gradients and film mulching on morphological characteristics of peanut root system and yield in red soil under calcium deficiency were studied by setting pot experiment. The results showed that applying calcium fertilizer hardly affected the root biomass of peanut, but film mulching had a significant effect on root biomass, total root length, volume and surface area. Under open field cultivation total root length, surface area and volume of Ca50 treatment were 49.48%, 39.07% and 27.91% higher than that of Ca0 treatment, respectively. The ratio of root length density distribution in 0-20 cm layer of Ca25 and Ca50 treatment was 13.53% and 17.22% higher than that of Ca0 treatment. Under the same calcium fertilizer treatment, the root surface area and volume in different soil layers of plastic film cultivation were higher than those in open field cultivation. The order of the surface area and volume of roots in 0-20 cm and 40 cm soil layers in different calcium treatments was Ca50> Ca25> Ca0, and the surface area and volume of roots of Ca50 treatment were higher 54.99% and 35.97% than those of Ca0 treatment, respectively. There was a highly significant positive correlation among total root length, root length or surface area in 0-20 cm soil layer and yield, and there was a significant positive correlation between total root surface area and yield. The main contribution of calcium fertilizer and plastic film to peanut was to promote the development of inner root and capillary root in 0-20 cm soil layer, so that the root system was more developed, and the surface area and volume of root in different soil layers increased, which further improved the ability on absorption and utilization of deep soil water, nutrients and resulted higher plant population quality, yield and harvest index.
To explore effect of interaction between biochar-based fertilizer and water stress on peanut production, potted experiment was conducted to investigate the effect of different levels of biochar-based fertilizer (BF0, 0kg/hm2; BF1, 750 kg/hm2; BF2, 1500 kg/hm2) and water stress (W0, normal irrigation, 70%-75% of field capacity; W1, moderate water stress, 60%-65% of field capacity; W2, severe water stress, 50%-55% of field capacity) on physiological characteristics and yield of peanut. The results showed that, compared with BF0, BF1 and BF2 treatments improved leaf growth, sustained higher chlorophyll content and net photosynthetic rate, increased shoot and root dry matter accumulation and thereby lead to higher yield. In addition to W2, W0 and W1 treatment both increased the net photosynthetic rate, dry matter accumulation and yield. Under moderate water stress, shoot dry matter accumulation and yield under biochar-based fertilizer application rate of 750 kg/hm2 and 1 500 kg/hm2 were increased by 52.2% and 39.9%, 62.0% and 42.5%, respectively, when compared to treatment without biochar-based fertilizer addition. And the dry matter accumulation was highly and positively correlated with peanut yield at pod filling stage.
A five-year field study with two factorial design was conducted to investigate the effects of different soil tillage (rotary tillage, RT and no tillage, NT) and wheat straw returning (no straw returning, M0; half straw returning, M1; total straw returning, M2) on soybean yield and soil physicochemical properties under wheat-soybean double cropping system in Yellow-Huai-Hai Rivers Valley (Xuzhou). The results showed that compared with rotary tillage, no tillage had no effects on soybean yield and the concentrations of organic matter and available K in soil, but resulted in lower NO3--N, and higher NH4+-N content in soil, and an increased available P and soil bulk density during sowing to V6. Compared to no straw returning, pod number per plant, 100-seed weight, yield and the content of soil organic matter and available K were increased; and the content of soil NO3--N and NH4+-N, soil bulk density during R1 to R7 were decreased in the treatment of wheat straw returning. Moreover, no tillage and straw returning decreased soil pH and soil temperature during sowing to R1, and increased soil humidity during sowing to R1. Pod number per plant, 100-seed weight, yield, soil organic matter content, soil pH and soil NO3--N content in no tillage treatment were higher than those of rotary tillage treatment under no straw returning. However, compared with rotary tillage, no tillage increased soil humidity, the content of soil organic matter and available P, and decreased soil temperature, the content of NO3--N and NH4+-N content under straw returning. There was no significant difference in soybean yield and soil available K content between rotary tillage and no tillage treatments. Compared with NS + M1, NS + M2 decreased soil pH and soil temperature, and increased soil humidity and soil organic matter content. Hence, no tillage in combination with straw returning, especially total straw returning, should be used and more nitrogen fertilizer should be applied, in Yellow-Huai-Hai River Valley.
A Trichoderma spp. strain TM was isolated from the sclerotia of Sclerotinia sclerotiorum in peanut in Shuangcheng, Heilongjiang Province. Based on the methods of morphological identification and molecular biology analysis, the strain was identified as Trichoderma koningiopsis. The results of dual culture demonstrated that T. koningiopsis strain TM had a significant inhibition activity on S. sclerotiorum in peanut. The colony of TM strain grew over that of S. sclerotiorum, and inhibited sclerotia formation of S. sclerotiorum or degraded sclerotia. The result of light microscopy observation noted that TM could grow paralleling, coiling around or penetrating the hyphae of S. slerotiorum, resulting in inhibiting mycelium growth or degrading the hyphae. The antimicrobial activity of TM strain against S. sclerotiorum demonstrated that the mycelium growth of S. sclerotiorum could be efficiently suppressed by non-volatile antibiotic substance of TM. In conclusion, the biocontrol mechanism of T. koningiopsis strain TM against S. sclerotiorum in peanut was that TM could produce antimicrobial metabolites in the mycoparasitism process, resulting in degrading the hyphae and inhibiting sclerotia formation of S. sclerotiorum.
To improve the yield of silage rape after spring wheat, field experiments were conducted on fertilization effect, yields response and principal of NPK uptake by silage rape. Results showed that rape yield in the treatment of combined application of N, P and K fertilizers were 63.9%, 28.3% and 12.7% higher than that of PK, NK and NP treatments respectively. The fertilization effect was in the order of N>P>K. The nutrients uptake for producing 1 ton silage rape were 47.3kg (K2O), 27.4kg (N) and 9.2kg (P2O5). N application greatly influenced plant height and organic matter, followed by P and K. Nutrients accumulation was on "slow-fast-slow" curves. Rape had the highest uptake under NPK treatment, and the lowest uptake under PK treatment. N, P2O5 and K2O accumulation peaks appeared on the 44-49th, 47-55th and 43-51st days after seeding, which indicated the critical periods of NPK application for high yield. Each treatment in wheat/silage rape multiple cropping had no effect for the following crop and had no effect on soil organic matter and alkali-hydrolyzale nitrogen content after two-year experiments of wheat/silage rape multiple cropping. The NPK treatment of wheat/silage rape multiple cropping could increase the soil Olsen P content but decrease the available K content.
Phytophthora root rot is one of the most important diseases of soybean. Reactions of 77 soybean cultivars (lines) to 8 strains of P. sojae were identified by using a modified hypocotyls inoculation technique. Phytophthora resistance genes in the cultivars (lines) were postulated by comparisons of the reaction types of differential soybean cultivars or lines. 24 reaction types occurred in 77 cultivars (lines). Andou1498 was the only cultivar with RRRRRRRR reaction type among 77 soybean cultivars (lines). This paper would provide a technical support for the large-scale popularization of Andou1498 in the epidemic area of Phytophthora root rot and provide a theoretical base for soybean breeding as a new germplasm resource resistant to Phytophthora root rot.
20 samples of hemp seed oil were selected and their composition and the contents of fatty acids, triglycerides, phytosterols and tocopherols in the oils were determined. Hemp seed oil mainly contained 5 kinds of fatty acids, palmitic acid, stearic acid, oleic acid, linoleic acid and linolenic acid. The unsaturated fatty acid content was 89.4% ~ 91.8%, polyunsaturated fatty acid content was 73.8% ~ 80.5%. Hemp seed oil contained 28 kinds of triglyceride structure, the 3 highest contents of triglyceride structure in hemp seed oil were trilinolein (LLL), dilinoleic acid linolenic acid glyceride (LLLn), Dilinoleic acid oleic acid glyceride (OLL). Hemp seed oil contained 6 kinds of phytosterols of campesterol, stigmasterol, beta sitosterol, fucosterol, alpha -1 sitosterol and 9,19-cyclolanost-24-en-3-ol. The total phytosterols content was 92.5-590.4 mg/100g. Beta sitosterol content was the highest, accounted for about 60% of the total content; Hemp seed oil contained α-, β, γ-, ζ-tocopherol. The total tocopherols ranged from 27.7 to 91.2 mg/100g. Gamma tocopherol content was the highest, accounted for about 90% of the total content. According to clustering analysis on the basis of various nutrients content, the 20 origins of hemp seed oil could be divided into 5 types.
