Deep transcriptome sequencing allows for the acquisition of large-scale microsatellite information, and it is especially useful for genetic diversity analysis and mapping in plants without reference genome sequences. In this study, a total of 14,004 simple sequence repeats (SSRs) were mined from 10,511 unigenes screening of 63,608 nonredundant transcriptome unigenes in loquat (Eriobotrya japonica) with a frequency of 22 SSR loci distributed over 100 unigenes. Dinucleotide and trinucleotide repeat SSRs were dominant, accounting for 20.62%, and 42.1% of the total, respectively. Seventy primer pairs were designed from partial SSRs and used for polymerase chain reaction (PCR) amplification. Of these primer pairs, 54 exhibited amplification and 33 were polymorphic. The number of alleles at these loci ranged from two to 17, and the polymorphism information content values ranged from 0.24 to 0.89. We tested the transferability of 33 SSR polymorphic primer pairs in apple and pear, and the transferability rates in these two species were 90.9% and 87.9%, respectively. A high level of marker polymorphism was observed in apple [Malus ×domestica (66.7%)], whereas a low level was observed in pear [Pyrus sp. (51.5%)]. In addition, the PCR products from seven SSR primer pairs were selected for sequence analysis, and 89.2% of the fragments were found to contain SSRs. SSR motifs were conserved among loquat, apple, and pear. According to our sequencing results for real SSR loci, ≈12,490 SSR loci were present in these loquat unigenes. The cluster dendrogram showed a distinct separation into different groups for these three species, indicating that these SSR markers were useful in the evaluation of genetic relationships and diversity between and within the species of Maloideae in the Rosaceae. The results of our identified SSRs should be useful for genetic linkage map construction, quantitative trait locus mapping, and molecular marker-assisted breeding of loquat and related species.
Xiaoying Li, Hongxia Xu, Jianjun Feng and Junwei Chen
Ren-jun Feng, Li-li Zhang, Jing-yi Wang, Jin-mei Luo, Ming Peng, Jun-feng Qi, Yin-don Zhang and Li-fang Lu
Cold stress is one of the most important environmental factors affecting crop growth and agricultural production. Induced changes of gene expression and metabolism are critical for plants responding and acclimating to cold stress. Banana (Musa sp.) is one of the most important food crops in the tropical and subtropical countries of the world. Banana, which originated from tropical regions, is sensitive to cold, which can result in serious losses in commercial banana production. To investigate the response of the banana to cold stress conditions, changes in protein expression were analyzed using a comparative proteomics approach. ‘Brazil’ banana (Musa acuminata AAA group) is a common banana cultivar in southern China. ‘Brazil’ banana plantlets were exposed to 5 °C for 24 hours and then total crude protein was extracted from treatment and control leaves by phenol extraction, separated with two-dimensional gel electrophoresis, and subsequently identified by mass spectrometry (MS). Out of the more than 400 protein spots reproducibly detected, only 41 protein spots exhibited a change in intensity by at least 2-fold, with 26 proteins increasing and 15 proteins decreasing expression. Of these, 28 differentially expressed proteins were identified by MS. The identified proteins, including well-known and novel cold-responsive proteins, are involved in several cellular processes, including antioxidation and antipathogen, photosynthesis, chaperones, protein synthesis, signal transduction, energy metabolism, and other cellular functions. Proteins related to antioxidation, pathogen resistance, molecular chaperones, and energy metabolism were up-regulated, and proteins related to ethylene synthesis, protein synthesis, and epigenetic modification were down-regulated in response to cold temperature treatment. The banana plantlets incubated at cold temperatures demonstrated major changes in increased reactive oxygen species (ROS) scavenging, defense against diseases, and energy supply. Increased antioxidation capability in banana was also discovered in plantain, which has greater cold tolerance than banana in response to cold stress conditions. Therefore, we hypothesized that an increased antioxidation ability could be a common characteristic of banana and plantain in response to cold stress conditions. These findings may provide a better understanding of the physiological processes of banana in response to cold stress conditions.
Xia Ye, Xianbo Zheng, Dehua Zhai, Wen Song, Bin Tan, Jidong Li and Jiancan Feng
Ethylene is important during the berry development and in the last stages of rachis development or rachis senescence. Since grapes develop in a cluster that comprises both the fruit berry and the nonfruit rachis, we measured the release of ethylene from both tissues. Detached berries from Vitis vinifera ‘Ruby Seedless’ and ‘Thompson Seedless’ showed that ethylene release peaks at the beginning of berry development and at veraison. Ethylene production in the rachis was higher than that in the berry and had an obvious peak before harvest in ‘Thompson Seedless’. In both cultivars, ethephon treatment induced ethylene production in the rachis but not in the berry. Expression of 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) and ACC oxidase (ACO) genes showed diverse temporal and spatial patterns in ‘Thompson Seedless’ and ‘Ruby Seedless’. For most gene family members, the low ACS expression levels were observed in berry and rachis. Expression levels of most of the ACS and ACO genes did not correlate with ethylene released in the same organ. The transcriptional level of VvACS1 did correlate with ethylene evolution in rachis of ‘Thompson Seedless’ during berry development and storage, which suggested that VvACS1 may have important roles in rachis senescence. In berries of ‘Thompson Seedless’ and ‘Ruby Seedless’, the transcriptional levels of VvACO1, VvACS2, and VvACS6 coincided with ethylene production, indicating possible roles in berry development. Expression of VvACS2–VvACO9 and VvACO1–VvACO3 was not consistent with ethylene production during storage or in response to ethephon treatment, which suggests that the expression of ACS and ACO was affected by other stress factors after harvest.
Joseph N. Wolukau, Xiao-Hui Zhou, Ying Li, Yong-Bin Zhang and Jin-Feng Chen
Gummy stem blight incited by the fungus Didymella bryoniae is a major disease of melons worldwide. The objectives of the present study were to critically evaluate melon (Cucumis melo L.) germplasm for resistance to D. bryoniae and to characterize the genetics of resistance in the resistant accessions. Two hundred sources of germplasm (plant introduction accessions, cultivars, breeding lines, landraces, and wild relatives) were screened against a single highly virulent isolate (IS25) of D. bryoniae in a plastic tunnel. The genetics of resistance to D. bryoniae was studied in three crosses between plant introductions 157076, 420145, and 323498, resistant parents that were fairly adapted (flowering, fruiting, powdery mildew tolerance) to Nanjing conditions, and plant introductions 268227, 136170, and NSL 30032 susceptible parents, respectively. Six populations of each cross (susceptible parent, resistant parent, F1, F2, the two reciprocal backcrosses) were analyzed for their responses to D. bryoniae. Seedlings in both studies were inoculated with a spore suspension (5 × 105 spores/mL−1) of D. bryoniae at the four to six true-leaf stages and assessed for leaf and stem damage at 7, 14, and 21 d postinoculation. Results of germplasm screening indicated most germplasms reported as resistant elsewhere were confirmed resistant under our conditions. However, some plant introductions identified as highly resistant elsewhere were susceptible under our conditions, the most interesting being plant introduction 482399. This plant introduction that was considered resistant was highly susceptible in our study. We also identified other sources of resistance not reported previously, for example, JF1; a wild Cucumis from the highlands of Kenya was rated highly resistant. Analysis of segregation of F1, F2, and backcross generations of the three crosses indicated that each of the three plant introductions carry a single dominant gene for resistance to the D. bryoniae.
Xiaoyuan Feng, Baogang Wang, Wensheng Li, Lei Shi, Jiankang Cao and Weibo Jiang
Preharvest application of Phellodendron bark (Phellodendron chinese Schneid) extract (PBE) on brown rot and postharvest quality of peach [Prunus persica (L.) Batsch var. platycarpa (Decne.) L.H. Bailey] was investigated. PBE at 0.8, 1.6, and 3.2 mg·mL−1 totally inhibited conidial germination, mycelial growth, and sporulation, respectively, of Monilinia fructicola in vitro. Preharvest PBE treatment at 21.0 mg·mL−1 at 0, 30, 60, and 90 days after full bloom controlled brown rot caused by M. fructicola on peach fruit after harvest and reduced disease incidence and lesion diameter by 37% and 61%, respectively, than those of the control 96 h after inoculation in in vivo experiments. The results from field experiments were consistent during a 3-year period. Fruit from PBE-treated trees showed higher activities of defense enzymes, including peroxidase, phenylalanine ammonia-lyase, chitinase, and β-1,3-glucanase, compared with those of the control during storage. PBE also delayed softening and loss of titratable acidity and inhibited flesh browning during storage. Total soluble solid contents were unaffected by treatment. The results indicate that preharvest application of PBE may be an alternative for controlling brown rot of peach fruit.
Weisheng Liu, Dongcheng Liu, Aimin Zhang, Chenjing Feng, Jianmin Yang, Jaeho Yoon and Shaohua Li
Inter-simple sequence repeat (ISSR) markers were used to evaluate genetic similarity and interrelationship among 104 plum (Prunus L. spp.) and related accessions from the Chinese National Germplasm Repository for Plums and Apricots and the Tianshan Germplasm Repository for Wild Fruit Resources, including six plum species (Prunus salicina Lindl., Prunus simonii Carr., Prunus ussuriensis Kov. et Kost., Prunus domestica L., Prunus cerasifera Ehrh., and Prunus spinosa L.), two related species [apricot (Prunus armeniaca L.) and nanking cherry (Prunus tomentosa Thunb.)], eight putative hybrids between plum and apricot (plumcot), and six accessions of wild European plum (P. domestica). Out of the 42 ISSR primers, 12 were selected, which generated 103 markers in total, 99 of which were polymorphic. Possible accession-specific ISSR bands or patterns were also found. Some possible synonyms or homonyms were clarified or discussed, and closely related accessions such as bud mutants were discriminated. Based on the unweighted pair group method with arithmetic mean (UPGMA) analysis and principal coordinate analysis (PCoA) using the Jaccard coefficient, two different dendrograms were constructed—one including accessions grouped by species and one with all 104 accessions—and a two-dimensional plot was obtained. Three groups were formed in both dendrograms and PCoA plot: Group I including apricot (‘Yinxiangbai’) and plumcot types; Group II containing Asia-originated diploid species [e.g., P. cerasifera, P. ussuriensis, P. tomentosa, and Chinese plum-types (i.e., P. salicina and its hybrids)]; and Group III involving European-origin polyploid species (e.g., P. spinosa and P. domestica) and recently found wild European plum accessions in China. The dendrogram with accessions grouped by species implied that 1) plumcot types had closer relatedness with apricot than with plum; 2) P. simonii should be a variant of P. salicina while P. ussuriensis an independent species; 3) P. domestica was more closely related to P. spinosa than to P. cerasifera. Two accessions of European plum (‘89-7-3’ and ‘Wanhei’) were clustered into outgroups in the dendrogram with all 104 accessions, which could been grouped within Group III in the PCoA plot. The distribution of both European plum and Chinese plum-types across respective groups did not reflect the geographic origins. The present study also further confirmed that the wild plants found in Xinjiang of China were P. domestica.
Kai Zhao, Feng Zhang, Yi Yang, Yue Ma, Yuexue Liu, He Li, Hongyan Dai and Zhihong Zhang
GA20-oxidase (GA20-ox) is a key enzyme involved in the biosynthesis of gibberellic acid (GA). To investigate its role in plant growth and development, we suppressed MdGA20-ox gene expression in apple (Malus domestica cv. Hanfu) plants by RNA interference (RNAi). After 20 weeks of growth in the greenhouse, significant phenotype differences were observed between transgenic lines and the nontransgenic control. Suppression of MdGA20-ox gene expression resulted in lower plant height, shorter internode length, and higher number of nodes compared with the nontransgenic control. The expression of MdGA20-ox in transgenic plants was significantly suppressed, and the active GA content in transgenic lines was lower than that in the nontransgenic control. These results demonstrated that the MdGA20-ox gene plays an important role in vegetative growth, and therefore it is possible to develop dwarfed or compact scion apple cultivars by MdGA20-ox gene silencing.
Wenlei Guo, Li Feng, Dandan Wu, Chun Zhang and Xingshan Tian
Widespread herbicide-resistant weeds and severe insect pest infestations pose a challenge to the preplant pest management (PPPM) strategy currently in use in leaf vegetable fields in southern China. The aim of this study was to develop a new weed and insect control method for use before planting leaf vegetables in southern China. Two flaming machines (a tractor mounted and a trolley flaming machine) were designed, and their efficacies for the control of insect and weed pests were evaluated and compared in two field trials. With liquefied petroleum gas (LPG) at 101 kg·ha−1, flaming machines reduced plant numbers by 86.7% to 98.8% 2 days after treatment (DAT), which was equal to or higher than the reduction after application of paraquat at 900 g·ha−1. Some weed species, especially awnless barnyard grass (Echinochloa colona) and goosegrass (Eleusine indica), regrew at 7 DAT, resulting in a decrease in control efficacy. Flaming machines also reduced the number of diamondback moth (Plutella xylostella) larvae by 83.0% to 88.2% and the number of adult striped flea beetles (Phyllotreta striolata) by 64.9% to 80.9%. This is the first report on flaming treatment in China to show that this method is a promising alternative to chemical pesticides for PPPM in leaf vegetable fields.
Ting-Ting Li, Zhi-Rong Li, Kang-Di Hu, Lan-Ying Hu, Xiao-Yan Chen, Yan-Hong Li, Ying Yang, Feng Yang and Hua Zhang
Kiwifruit (Actinidia deliciosa) is a typical climacteric fruit, and its ripening is closely associated with ethylene. In this study, we present evidence that H2S alleviated ethylene-induced ripening and senescence of kiwifruit. Kiwifruit were fumigated with ethylene released from 0.4 g·L−1 ethephon solution or H2S with 1 mm sodium hydrosulfide (NaHS) as the donor or in combination. Fumigation with ethylene was found to accelerate kiwifruit ripening and H2S treatment effectively alleviated ethylene-induced fruit softening in parallel with attenuated activity of polygalacturonase (PG) and amylase. Ethylene + H2S treatment also maintained higher levels of ascorbic acid, titratable acid, starch, soluble protein, and reducing sugar compared with ethylene group, whereas suppressed the increase in chlorophyll and carotenoid. Kiwifruit ripening and senescence under ethylene treatment was accompanied by elevation in reactive oxygen species (ROS) levels, including H2O2 and superoxide anion and malondialdehyde (MDA), but combined treatment of ethylene plus H2S alleviated oxidative stress in fruit. Furthermore, the activities of antioxidative enzymes catalase (CAT) and ascorbate peroxidase (APX) were increased by ethylene + H2S treatment in comparison with ethylene alone, whereas the activities of lipoxygenase (LOX) and polyphenol oxidase (PPO) were attenuated by H2S treatment. Further investigations showed that H2S repressed the expression of ethylene synthesis-related genes AdSAM, AdACS1, AdACS2, AdACO2, and AdACO3 and cysteine protease genes, such as AdCP1 and AdCP3. Taken together, our findings suggest that H2S alleviates kiwifruit ripening and senescence by antagonizing the effect of ethylene through reduction of oxidative stress and inhibition of ethylene synthesis pathway.
Feng Gao, Arvind H. Hirani, Jun Liu, Zheng Liu, Guohua Fu, Chunren Wu, Peter B.E. McVetty and Genyi Li
There are various clubroot pathogen (Plasmodiophora brassicae) resistance genes within Brassica species with european turnip (B. rapa ssp. rapifera) being identified as potentially the best source of resistance for the development of clubroot-resistant cultivars in chinese cabbage (B. rapa ssp. pekinensis). To use clubroot resistance genes effectively, it is necessary to map these genes so that molecular markers inside or closely linked to these resistance genes can be developed. Using molecular marker-assisted selection, the clubroot resistance genes can be effectively transferred from cultivar to cultivar and from species to species. In this report, one clubroot resistance locus was mapped on linkage group A3 using five segregating populations developed from five chinese cabbage cultivars, suggesting that all the five cultivars shared the same clubroot resistance locus. Furthermore, one of these five chinese cabbage cultivars was used to develop a large segregating population to fine-map this clubroot resistance locus to a 187-kilobp chromosomal region. Molecular markers that are closely linked to the mapped clubroot resistance locus have been developed that can be used for marker-assisted selection in chinese cabbage and canola/rapeseed (B. rapa and B. napus) breeding programs.