Synergistic effects of carbon cycle metabolism and photosynthesis in Chinese cabbage under salt stress

Chinese cabbage(Brassica rapa ssp. pekinensis) has a long cultivation history and is one of the vegetable crops with the largest cultivation area in China. However, salt stress severely damages photosynthesis and hormone metabolism, nutritional balances, and results in ion toxicity in plants. To better understand the mechanisms of salt-induced growth inhibition in Chinese cabbage, RNA-seq and physiological index determination were conducted to explore the impacts of salt stress on carbon cycle metabolism and photosynthesis in Chinese cabbage. Here, we found that the number of thylakoids and grana lamellae and the content of starch granules and chlorophyll in the leaves of Chinese cabbage under salt stress showed a time-dependent response, first increasing and then decreasing. Chinese cabbage increased the transcript levels of genes related to the photosynthetic apparatus and carbon metabolism under salt stress, probably in an attempt to alleviate damage to the photosynthetic system and enhance CO_(2) fixation and energy metabolism. The transcription of genes related to starch and sucrose synthesis and degradation were also enhanced;this might have been an attempt to maintain intracellular osmotic pressure by increasing soluble sugar concentrations. Soluble sugars could also be used as potential reactive oxygen species(ROS) scavengers, in concert with peroxidase(POD)enzymes, to eliminate ROS that accumulate during metabolic processes. Our study characterizes the synergistic response network of carbon metabolism and photosynthesis under salt stress.

Fine mapping and cloning of the sterility gene Bra2Ms in nonheading Chinese cabbage(Brassica rapa ssp.chinensis)

The application of a male-sterile line is an ideal approach for hybrid seed production in non-heading Chinese cabbage(Brassica rapa ssp.chinensis).However,the molecular mechanisms underlying male sterility in B.rapa are still largely unclear.We previously obtained the natural male sterile line WS24-3 of non-heading Chinese cabbage and located the male sterile locus,Bra2Ms,on the A2 chromosome.Cytological observations revealed that the male sterility of WS24-3 resulted from disruption of the meiosis process during pollen formation.Fine mapping of Bra2Ms delimited the locus within a physical distance of about 129 kb on the A2 chromosome of B.rapa.The Bra039753 gene encodes a plant homeodomain(PHD)-finger protein and is considered a potential candidate gene for Bra2Ms.Bra039753 was significantly downregulated in sterile line WS24-3 compared to the fertile line at the meiotic anther stage.Sequence analysis of Bra039753 identified a 369 bp fragment insertion in the first exon in male sterile plants,which led to an amino acid insertion in the Bra039753 protein.In addition,the 369 bp fragment insertion was found to cosegregate with the male sterility trait.This study identified a novel locus related to male sterility in non-heading Chinese cabbage,and the molecular marker obtained in this study will be beneficial for the marker-assisted selection of excellent sterile lines in non-heading Chinese cabbage and other Brassica crops.

Physiological and transcriptome analyses of Chinese cabbage in response to drought stress

Chinese cabbage is an important leafy vegetable crop with high water demand and susceptibility to drought stress.To explore the molecular mechanisms underlying the response to drought,we performed a transcriptome analysis of drought-tolerant and-sensitive Chinese cabbage genotypes under drought stress,and uncovered core drought-responsive genes and key signaling pathways.A co-expression network was constructed by a weighted gene coexpression network analysis(WGCNA)and candidate hub genes involved in drought tolerance were identified.Furthermore,abscisic acid(ABA)biosynthesis and signaling pathways and their drought responses in Chinese cabbage leaves were systemically explored.We also found that drought treatment increased the antioxidant enzyme activities and glucosinolate contents significantly.These results substantially enhance our understanding of the molecular mechanisms underlying drought responses in Chinese cabbage.

Defect in an immune regulator gene BrSRFR1 leads to premature leaf senescence in Chinese cabbage

Leaf senescence is the final stage of leaf development, where the nutrients and energy of senescent leaves are redistributed to developing tissues or organs for plant growth, reproduction, and defense. Outer leaves are photosynthetic organs that usually senesce at the late heading stage in Chinese cabbage, and premature leaf senescence often reduces leafy head yield and quality. In this study, 11 premature leaf senescence mutants were screened from an ethyl methanesulfonate-mutagenized population of the double haploid line ‘FT' in Chinese cabbage. At the early heading stage, the mutants exhibited edge yellowing within its outer leaves, and at the mature stage, its leafy head weight decreased significantly. Genetic analysis revealed that the mutated trait of all 11 mutants corresponds to single gene recessive inheritance. Semi-diallel cross tests showed that 5 of the 11 were allelic mutants. MutMap and Kompetitive Allele Specific PCR genotyping revealed that BraA01g001400.3C was the candidate gene, which is orthologous of Arabidopsis SUPPRESSOR OF rps4-RLD 1, encoding an immune regulator, so we named it as BrSRFR1. All the BrSRFR1 in the five allelic mutants exhibited single nucleotide polymorphisms at different positions on their exons and led to premature translation termination, which confirmed that defect in BrSRFR1 led to premature leaf senescence. These results verify the role of Br SRFR1 on leaf senescence and provide a new insight into the mechanisms of leaf senescence in Chinese cabbage, which reveals a novel function of SRFR1 in plant development.

Fast,simple,efficient Agrobacterium rhizogenes-mediated transformation system to non-heading Chinese cabbage with transgenic roots

Non-heading Chinese cabbage, a variety of Brassica campestris, is an important vegetable crop in the Yangtze River Basin of China. However,the immaturity of its stable transformation system and its low transformation efficiency limit gene function research on non-heading Chinese cabbage. Agrobacterium rhizogenes-mediated(ARM) transgenic technology is a rapid and effective transformation method that has not yet been established for non-heading Chinese cabbage plants. Here, we optimized conventional ARM approaches(one-step and two-step transformation methods) suitable for living non-heading Chinese cabbage plants in nonsterile environments. Transgenic roots in composite non-heading Chinese cabbage plants were identified using phenotypic detection, fluorescence observation, and PCR analysis. The transformation efficiency of a two-step method on four five-day-old non-heading Chinese cabbage seedlings(Suzhouqing, Huangmeigui, Wuyueman, and Sijiu Caixin) was 43.33%-51.09%, whereas using the stout hypocotyl resulted in a transformation efficiency of 54.88% for the 30-day-old Sijiu Caixin.The one-step method outperformed the two-step method;the transformation efficiency of different varieties was above 60%, and both methods can be used to obtain transgenic roots for functional studies within one month. Finally, optimized ARM transformation methods can easily,quickly, and effectively produce composite non-heading Chinese cabbage plants with transgenic roots, providing a reliable foundation for gene function research and non-heading Chinese cabbage genetic improvement breeding.

Fast tracking alien gene discovery by molecular markers in a late flowering Chinese cabbage-cabbage translocation line‘AT7–4'

Flowering time is an important agronomic trait of Chinese cabbage with late flowering being a primary breeding objective.In our previous work,we obtained Chinese cabbage-cabbage translocation lines that contained several beneficial cabbage genes.Cabbage-specific molecular markers show that these genes were coming from chromosome C01 of cabbage.In this study,we investigated the inheritance of flowering time in a couple of translocation lines and analyzed the transmission rate of molecular markers in the offspring.Consequently,we obtained the late flowering Chinese cabbage-cabbage translocation line‘AT7–4’in which the flowering time was later than that of‘85–1’by about 7 days under 4-week vernalization.Based on previous studies of the genomes of Chinese cabbage and cabbage,we located the cabbage-specific molecular markers that were closely linked at the top of the chromosome A01 in the F2mapping population generated by self-crossing F1s derived from a cross between the translocation line‘AT7–4’and Chinese cabbage‘14–36’.Five flowering-related genes in the alien fragment were found by functional annotation and their molecular markers were developed.This study lays the foundation for the future improvement of Chinese cabbage varieties using A-C translocation lines.

Blue light induces leaf color change by modulating carotenoid metabolites in orange-head Chinese cabbage(Brassica rapa L.ssp. pekinensis)

Carotenoids are involved in the formation of plant leaf color as well as photosystem photoprotection. This study showed that blue light significantly induced up-regulation of the total carotenoid content in the inner leaves of orange-head Chinese cabbage(OHCC). Furthermore, the transcriptomic analysis revealed that blue light treatment induced upregulation of genes in photosynthesis(BrHY5-2, BrCOP1 and BrDET1) and the methylerythritol 4-phosphate pathways(BrGGPS, BrDXS and BrHDR) upstream of the carotenoid metabolic pathway. Carotenoid metabolomic analysis revealed that the accumulation of several orange and red carotenoids(lycopene, zeaxanthin, β-carotene, lutein, and β-cryptoxanthin) after blue light treatment contributed to the deepening of the leaf coloration, suggesting that short-term blue light treatment could be used to boost nutritional quality. The light signal gene BrHY5-2 participated in the blue light-induced transcriptional regulation of carotenoid biosynthesis in OHCC. Overexpression of BrHY5-2 in Arabidopsis significantly increased the total carotenoid content and the sensitivity to blue light. The above findings revealed new insights about blue-light-induced carotenoid synthesis and accumulation in OHCC lines. They suggested a new engineering approach to increase the nutritional value of vegetables.

Comprehensive analysis of the full-length transcripts and alternative splicing involved in clubroot resistance in Chinese cabbage

Chinese cabbage is an economically important Brassica vegetable worldwide, and clubroot, which is caused by the soilborne protist plant pathogen Plasmodiophora brassicae is regarded as a destructive disease to Brassica crops. Previous studies on the gene transcripts related to Chinese cabbage resistance to clubroot mainly employed RNA-seq technology,although it cannot provide accurate transcript assembly and structural information. In this study, PacBio RS II SMRT sequencing was used to generate full-length transcriptomes of mixed roots at 0, 2, 5, 8, 13, and 22 days after P. brassicae infection in the clubroot-resistant line DH40R. Overall, 39 376 high-quality isoforms and 26 270 open reading frames(ORFs) were identified from the SMRT sequencing data. Additionally, 426 annotated long noncoding RNAs(lncRNAs),56 transcription factor(TF) families, 1 883 genes with poly(A) sites and 1 691 alternative splicing(AS) events were identified. Furthermore, 1 201 of the genes had at least one AS event in DH40R. A comparison with RNA-seq data revealed six differentially expressed AS genes(one for disease resistance and five for defensive response) that are potentially involved in P. brassicae resistance. The results of this study provide valuable resources for basic research on clubroot resistance in Chinese cabbage.

Effect of Foliar Application of Amino Acid on the Quality and Enzyme Activity of Flowering Chinese Cabbage(Brassica parachinensis Bailey)

A pot experiment was conducted to study the influences of foliar application of glycine,alanine,lysine,and glutamic acid in 200 mg/kg or 500 mg/kg upon the quality and enzyme activity of flowering Chinese cabbage（Brassica parachinensis Bailey）.The results showed that all the application of these four amino acids could increase the yield of flowering Chinese cabbage,significantly raise the content of soluble sugar,and reduce the accumulation of nitrate.The applications of three other amino acids except alanine can increase the content of soluble proteins and decrease the accumulation of oxalic acid.However,the application of amino acid has insignificant influences on the SPAD number of chlorophyll,and causes the decrease of Vitamin C content.Meanwhile,the application of amino acid can improve the activity of nitrate reductase（NR） and glutamate dehydrogenase（GDH） as well.It shows that the application of amino acid is beneficial to improve ammonia metabolism,reduce the accumulation of nitrate and oxalic acid,increase the content of soluble sugar and soluble proteins,and improve the quality of flowering Chinese cabbage.

Investigation on the Sterility Changeover of Male Sterility Line CMS7311 in Heading Chinese Cabbage

Sterility changeover induced by low temperature in male sterility line CMS7311 of heading Chinese cabbage ( Brassica campestris L. ssp. penkinsis) was investigated under constant temperature, day/night alternating temperature in light-growth-incubator and outdoor natural day/night alternating temperature respectively. It showed that the sterility of CMS7311 has significant change with temperature. The constant temperature of 6 - 12 degreesC was suitable to induce the changeover of sterility (CGS) of CMS7311 ; the shortest treatment time for CGS induction at this temperature was 3 d. The significant value of sterility changeover of CMS7311 occurred at the constant temperature of 6 - 9 degreesC after treatment for 6 - 9 d. The intensity of CGS was stronger under constant temperature than under alternate temperature, but the shortest treatment time inducing CGS has no difference between them. When treated for 9 d under the day/night alternating temperature of 11/7 degreesC, its sterility could also turn to normal fertility (near to sterile grade I or zero). In normal condition, the interval from the end of temperature treatment to the beginning of CGS induction was 10 - 16 d. In the condition of natural day/night alternating temperature, the lower limit of temperature for effective CGS induction was at the daily mean temperature ( DMT) of 3. 5 - 6. 8 degreesC, and the shortest treatment time which could induce the sterility changeover was also longer than that under constant temperature, being 11d at DMT of 1.5 - 3.1 degreesC. In addition, for the apical dominance, the flower bud on the first lateral branch showed the CGS 8 - 10 d later than the flower bud on the main stem under the same temperature with the same days of treatment. The intensity of CGS of CMS7311 on the first lateral branch was also weaker than the flower bud on the main stem.

Effects of Hypergravity on Physiological and Biochemical Characteristics of Chinese Cabbage

The Chinese cabbage seed used in this study was Xinfengkang 90. The tested seeds were treated with 1 000×g, 2 000×g and 4 000×g of hypergravity for 20, 40 and 60 min respectively. The Chinese cabbage leaves were col ected at the early seedling stage for the determination of free proline content, POD activity and SOD activity. The results showed the free proline content was lowest in the seeds treated with 40-min 2 000-g gravity, while it was highest for the 60-min 4 000-g gravity treatment. Compared to the control, al the hypergravity treatments inhibited the synthesis of free proline. The POD activity was highest in the seeds treated with 20-min 1 000-g gravity, while it was lowest in the seeds treated with 60-min 2 000-g gravity. Considering the SOD activity, the 40-min 1 000-g gravity treatment was highest, and the 60-min 4 000-g gravity treatment was lowest. lt was indicated that appropriate hypergravity treatment was conducive to increasing the SOD activity.

Directional Breeding of An Oval-ecotype Male Sterile Line of Chinese Cabbage(Brassica campestris L. ssp. pekinensis)

[Objective] The study aimed to explore the method for directional breeding of a male-sterile line in oval-ecotype Chinese cabbage. [Method] Based on ＂Multiple Allele Hypothesis of Genic Male Sterile Chinese Cabbage＂, an inbred line ＇06048＇ of oval ecotype was used as the receptor, and male fertile plant of ＇AB12＇ was used as the donor line. Crossing, backcross, selfing, testcross and sibling were ap- plied to transfer the multiple alleles under the directional genetic model. [Result] Segregation ratio of every generation was consistent with theoretical value. A new male sterile line with 100% male sterility and ＇06048＇ horticultural traits was ob- tained successfully, which accomplished the transfer of male sterile multiple allele and horticultural characters of receptor line at the same time. [Conclusion] The re- search verifies that the model of directional transfer is feasible, provides a theoreti- cal basis for the directional transfer of Chinese cabbage with other horticultural traits whose genotype is msms. The model can also be applied to other Brassica crops, to generate genetic male sterile lines with specific botanical traits and high-quality economic traits.

Evaluation of Chinese Cabbage Sensory Quality and Its Relationship with Contents of Main Nutrient Components

In order to establish a comprehensive evaluation system for quality of Chinese cabbage, 28 Chinese cabbage cultivars or inbreeds were selected as ex- perimental materials to identify the raw and cooked taste and determine the nutrient component contents, and the subjective factors （taste） and objective factors （nutrient component contents） influencing the sensory quality of Chinese cabbage were ana- lyzed by multiple regression and path analysis. The results showed that two regres- sion equations were obtained： yr （raw quality）=0.310 3＋0.254 0xl （juiciness）＋0.176 2 x2 （sweetness）＋0.221 6x3 （crispness）＋0.319 9x4 （flavor）; Yc （cooked quality）=0.204 4＋ 0.250 9 xs （residue）＋0.246 9x6 （cooked sweetness）＋0.182 5x7 （softness）＋0.323 lx8 （cooked flavor）. The path analysis results showed that among the sensory indices, the total coefficients of sweetness and juiciness were 0.445 3 and 0.427 6, respec- tively. It indicated they were the main factors affecting the raw quality of Chinese cab- bage, followed by flavor and crispness. Sweetness, with total coefficient of 0.515 2, showed the greatest effect on the cooked quality of Chinese cabbage, followed by residue, cooked flavor and softness. The effects of nutrient component contents on the sensory quality of Chinese cabbage were further analyzed, and the regression equa- tions were obtained： y, =-32.192 0＋0.389 3xw （moisture content）＋1.169 8x~ （soluble sugar content）; Yc = 7.497 1＋0.732 6xs - 5.668 8x, （organic acid content）-2.176 3x~ （crude fiber content）. The standardized regression coefficients of soluble sugar con- tent were all highest with the value of 0.580 7 in the raw quality regression equa- tion and 0.325 0 in the cooked quality one, indicating that among the six nutrient components, the soluble sugar content was the main factor affecting both raw and cooked quality of Chinese cabbage. This study will provide certain basis for quality evaluation, selection and breeding of high-quality Chinese cabbage.

Physiological and Biochemical Responses of Chinese Cabbage to La and Zn Stresses

[Objective] The aim of this study was to explore the physiological and biochemical responses of Chinese cabbage to La and Zn. [Method] The effects of La and Zn on seed germination and seedling growth were explored by tissue culture method. [Result] La and Zn had little effects on seed germination rate, but significantly inhibited the growth of root and seedling, decreased their fresh weight at higher concentration; the inhibition of La proved higher on Chinese cabbage. The contents of chlorophyll and soluble protein increased at lower concentrations of La and Zn, but decreased at higher concentrations. With increasing doses of La and Zn, the activities of CAT and POD rose gradually, while SOD activity decreased at lower doses and increased at higher doses. Both of La and Zn would promote Of. producing and MDA accumulation. [Conclusion] In higher concentration, La was more poisonous than Zn. It is clear that the poisonous mechanism of La is similar to that of Zn. Hence, La is possibly a kind of new pollutant.

Effect of Freshness Protection Package on Storage and Preservation of Chinese Cabbage

The effect of freshness protection package on Chinese cabbage placed in the self-made shelf in ventilated storage was investigated, and the physiological-qual-ity indexes change were determined during the storage period. The results showed that the losses of weight and vitamin C were significantly reduced. The accumula-tion of crude fiber was effectively inhibited. But protein content was maintained at a low level and the rotting more serious of Chinese cabbage at the end of the stor-age packaged with the fresh keeping bags.

Impacts of Irrigation with Reclaimed Water on Endophytic Bacteria in Chinese Cabbage in Urumqi City

[Objective] This study aimed to investigate the feasibility of agricultural irri- gation with reclaimed water in Urumqi City. [Method] Chinese cabbages were used as experimental materials and irrigated with control water, 50% reclaimed water and 100% reclaimed water, and then the number of endophytic bacteria in Chinese cab- bages was measured. [Result] Using 50% reclaimed water, the fresh weight of Chi- nese cabbage was improved by 68.94%; however, with the deepening internalization of exogenous microorganism internalization, the total number of endophytic bacteria and coliform flock in Chinese cabbages treated with 50% reclaimed water was sig- nificantly higher than the other two treatment groups （P〈0.05）. [Conclusion] Re- claimed water enhances both the weight of Chinese cabbage and the number of endophytic bacteria, which is unsuitable for irrigation of edible crops such as Chi- nese cabbage. Strengthening disinfection during the process of sewerage treatment could solve this problem.

Embryogenesis and Development of Isolated Microspore in Chinese Cabbage

[Objective] The aim was to observe embryogenesis and development of isolated microspores in Chinese cabbage. [Method] Chinese cabbage F 1 hybrids were used as the experimental materials, and optical microspore was employed to observe the embryogenesis and development of isolated microspores. [Result] Cells swelled after heat shock treatment, which was the critical factor of embryoid induction. Three pathways equal division, unequal division and germination of microspores were discovered to lead to the embryogenesis from isolated microspores after swelling. Microspore could grow directly to embryoid through germination path way. Equally divided microspores formed the original embryos after successive multiple equal divisions. Original embryos could develop into cotyledon-shaped embryos via globular, heart-shaped and torpedo-shaped embryos. The large one of the two cells from unequally divided microspores continued to divide and finally formed a polar embryoid. [Conclusion] The study will provide cytological basis for high induction frequency and embryoid of Chinese cabbage.

稀土对大白菜(Chinese cabbage)的效应机理及其应用技术

大白菜施用稀土增产效果达极显著水平,平均增产13.6%。主要原因是促进了根系发育,增加了根重和侧根数,扩大了根系垂直方向和水平方向的分布范围,为养分和水分的吸收和运转奠定了良好的基础,从而促进了大白菜地上部分的生长,增加了叶片数和叶面积。其主要机理是稀土提高了叶片中叶绿素的含量和酶的活性,从而增加了有机物质的合成;增强了对营养元素的吸收;增强了抗逆性,降低了发病率。

Disease Analysis and Pathogen Identification of Black Spot Disease of Chinese Cabbage [Alternaria brassicae (Berk.) Sace.] in Bijie City

[Objective] Disease analysis and pathogen identification of black spot disease of Chinese cabbage [Alternaria brassicae（Berk.） Sace.] in Bijie City of Guizhou Province was carried out,so as to raise the corresponding prevention measures.[Method] Pathogen was isolated from diseased leaves using tissue surface separation method for cultivation,microscopic examination,spore size determination and pathogen identification.[Result] Microscopic examination showed that the disease was caused by Alternaria sp.,the size of its spores was（15.0-38.5） μm×（7.5-13.5） μm.Combining with the characteristics of host and pathogen,the pathogen caused black spot disease of Chinese cabbage was identified to be Alternaria brassicicola.[Conclusion] A.brassicicola was the pathogenic fungi of black spot disease of Chinese cabbage in Bijie City of Guizhou Province,and damage caused by the disease could be reduced through agricultural measures and chemical control.

Breeding and Off-season Cultivation Techniques of a New Late-bolting Chinese Cabbage Variety Shiyuchunbao

Shiyuchunbao is a new hybrid of early-mid maturity spring Chinese cabbage with growth period of 60-65 d, flat-topped head and yellow inner leaves, characterized with high quality and good marketability. It is highly resistant to viral diseases, downy mildew and soft rot disease, and can be cultivated in Beijing, Hebai, Heilongjiang and Yunnan.