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Watermelon is an annual vegetable crop that suffers from various diseases during growth. Genetic engineering is an effective tool for improving plant disease resistance. This paper presents an effective transformation protocol designed for a female parent watermelon line mediated by Agrobacterium. In this protocol, 2 times MS Fe-EDTA were supplemented into Murashige and Skoog (MS) medium to suppress the chlorosis during watermelon explants culturing, and the optimal preculture time (5 days) and coculture time (5 days) were screened out to improve the frequency of shoots regeneration. Using our protocol, 17 putative transgenic plants expressing artificial microRNAs that target Cucumber mosaic virus (CMV) 2a/2b genes were recovered, and seven of them were succeeded in obtaining T1 generation seeds because of the technique difficulties of self-pollination of the transgenic watermelon lines. PCR and Southern blot analyses confirmed that the transferred fragment was successfully imported and integrated into the genome of three of these seven transgenic lines. DAS-ELISA assay indicated that the CMV can be detected in the nontransgenic line but not in the transgenic lines, and the transgenic lines displayed resistance to CMV infection.

The ascorbic acid (AsA) accumulation in plants is affected by various growth, development and environmental factors. The pH of growth media has been reported to affect the accumulation of AsA in plants. In this study, the impact of the pH on the AsA accumulation as well as its biosynthetic gene expression was analyzed in tomato (Solanum lycopersicum ‘Ailsa Craig’). The results indicated that with the increase in pH, total AsA content in tomato leaves decreased and redox status (AsA/DHA) was augmented. Similarly, most of the biosynthetic gene expression level decreased with the increment of pH and showed the negative correlation; only a few genes, e.g. , genes encoding GDP-Mannose 3′,5′-epimerase 1 (GME1), GDP- l -galactose-1-phosphate phosphorylase 1 (GGP1) and GGP2 showed positive correlation. Moreover, in situ H 2 O 2 was detected which accretion was escalated as the pH value surged from 5.0 to 7.0, in agreement to the decreased AsA level. The present study suggests that mild acidity may contribute to the boosted accumulation of AsA in tomato.

The global issue of heavy metal pollution causes concern on lotus root, a popular aquatic vegetable grown in sludge supposedly heavily contaminated. The relevance of heavy metals in soil and lotus roots in production remains unclear. In this work, four heavy metals, lead (Pb), cadmium (Cd), mercury (Hg) and arsenic (As), in lotus roots and ambient soil from main production areas of China were determined. The bioconcentration factor (BCF) of each heavy metal and correlation coefficient among heavy metals in soil and lotus roots were analyzed. Red soil (pH 6–7) in one location exceeded the maximum allowable limit of Cd by 91%, while soils in seven locations met the environmental requirements for producing safe lotus roots, including three red soils (pH 4–5), two moisture soils (pH 4–7), one cinnamon soil (pH 6–7), and one paddy soil (pH 6–7). Lotus roots were individually contaminated with Pb in two locations (red soil and paddy soil), As in two locations (red soil and cinnamon soil), and Hg in one location (red soil), exceeding the maximum allowable limit by 3–16%. Heavy metal levels of lotus roots produced in all investigated areas still met the safety standard as indicated by integrative pollution indexes (IPI). However, the close-to-limit IPI will certainly raise the safety concern for lotus root production. BCF of heavy metals decreased in an order of Cd (5.03–46.67%), Hg (2.42–7.20%), As (1.93–6.68%) and Pb (0.02–1.82%), showing lotus root's ability to accumulate metals varied with metal types. BCF and correlation coefficient showed that the lotus roots tended to accumulate heavy metals less efficiently from soil with higher levels of heavy metals, and vice versa. Heavy metal levels were positively correlated in soil but largely irrelevant in lotus roots. Thus, heavy metals in lotus roots were accumulated primarily independently from or partially negatively correlated with soil, complicating the enrichment mechanism of heavy metals in lotus roots. The possible causes of soil independent accumulation of heavy metals in lotus roots are discussed.

Reduced l -ascorbic acid can be oxidized by ascorbate peroxidase (APX) in ascorbate metabolism pathway. In present study, a mitochondrial APX (mitAPX) was cloned and suppressed by RNA interference in tomato via Agrobacterium -mediated transformation. The existence of the RNA interference T-DNA in transformed plants and their progenies were confirmed by polymerase chain reaction amplification and Southern blotting. Decreased mitAPX expression in homozygous transgenic lines was detected by semi-quantitative reverse transcriptase-polymerase chain reaction, accompanied by decreasing cytosolic APX activity by 0.4- to 0.8-fold of the untransformed control. The mitochondrial APX enzyme activity in RNAi lines decreased to 29–45% of the wild type plants. The down-regulated mitAPX expression and APX enzyme activity lead to 1.4- to 2.2-fold increase in ascorbic acid content in tomato fruits compared with wild-type control. The APX expression and APX enzyme activity were found to be correlated with increase in the amount of ascorbic acid in tomato fruits. This report demonstrates the modulation of ascorbic acid metabolic pathway by RNA interference method, and provides a feasible approach to improve ascorbic acid content in horticultural plants by suppressing genes related to ascorbic acid metabolism.

Purple flowering stalk (Brassica campestris ssp. chinensis var. purpurea Hort.) was generally regarded as a recalcitrant species for microspore culture in Brassica crops. Conditions for reliable induction of microspore embryogenesis were studied in 12 genotypes of purple flowering stalk. A treatment of short heat shock to microspore by incubating at 32 °C for 18 h was suitable for the survival of microspores, sustained cell divisions, and further induced embryogenesis. Subsequently, the reduced concentration of macro salts (1/2 NLN) provided an optimal condition for the development of embryoids. Under the optimized conditions for microspore embryoid development, 10 genotypes responded to microspore culture with the frequencies ranging from 2.7 to 70.5 embryoids per dish. However, regenerated plants were obtained from 9 genotypes, and more than 75% these regenerated plants were double haploid. This report establishes an efficient protocol for microspore culture and offers great potential for DH breeding in purple flowering stalk.

The amplification efficiency and identification diversity with gene-specific primers derived from S locus glycoprotein gene (SLG) and S locus receptor kinase gene (SRK) were compared, and the geographical distribution for S haplotypes was investigated by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) in 72 genotypes of 5 Brassica vegetables from China. The amplification efficiency and identification diversity with class I SRK primers were generally higher than that with class I SLG in most crops tested. Class I alleles were found in total 66 genotypes and they were classified into 16, 10, 7 and 10 groups for Chinese cabbage, purple flowering stalk, cauliflower and cabbage respectively. The number of amplification accessions and identification diversity using the primers of class II SLG and SRK were quite similar. Class II alleles were detected in 55 genotypes and further grouped into one type in mustard and three in other crops. The nucleotide sequences showed high similarity between identical S haplotypes determined by reciprocal pollination and PCR-RFLP tests. It demonstrated that the PCR-RFLP analysis was feasible for identification of S alleles, and SRK should be considered as a better marker for the identification of S haplotypes than SLG. The types of S haplotypes are highly diverse in Brassica vegetables from China. Nevertheless, they were geographically limited in some Brassica vegetables, so the exchange of germplasm resources should be enhanced for breeding.