The additive memory and healthspan enhancement effects by the combined treatment of mature silkworm powders and Korean angelica extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Silkworm (Bombyx mori) and Korean angelica (KoAg; Angelica gigas Nakai) have been widely used as traditional oriental medicines in Korea, China, and Japan to treat various diseases such as anemia, cold, diabetes, palsy, stroke, etc. Steamed and freeze-dried mature silkworm powder, also known as HongJam (HJ), and extracts of KoAg root (KoAgE) are currently sold in Korea as functional foods to improve memory, cognition, and liver functions. However, the molecular and pharmacological basis for the improvement of brain functions of HJ and KoAgE has not yet been elucidated. AIM OF STUDY: This study aimed to elucidate the molecular basis underlying the memory-enhancing effects of HJ and KoAgE and determine whether administration of HJ and KoAgE complexes (HJ+KoAgC) has additive memory and healthspan-enhancing effects. MATERIALS AND METHODS: The MCI mouse models generated by intraperitoneal injection of Scopolamine (Sco-IP) were orally administered with HJ and KoAgE alone or as complexes. Their memory-enhancing effects were examined on spatial, fear-aggravated, and social memories and compared with control or Donepezil (Dp) treatment. The activities of mitochondria complex (MitoCom) I-IV and acetylcholinesterase (AChE) and the amounts of ATP in the mouse brains were examined. The Drosophila model was used to investigate lifespan- and healthspan-promoting effects of HJ+KoAgC. RESULTS: Administration of HJ+KoAgC produced more memory-enhancing effects than administration of HJ or KoAgE alone or Dp. The increase in MitoCom I-IV activities and ATP amounts and the decrease in AChE activities in the mouse brains were the molecular basis for the memory enhancement. The greatest improvement in memory and mitochondrial function was observed when the mice were administered the 1:0.8 ratio of HJ+KoAgC. Administration of HJ+KoAgC to Drosophila prolonged the lifespan and the healthspan and increased the amounts of ATP. CONCLUSION: HJ+KoAgC had superior effects on memory improvement and healthspan extension by increasing mitochondrial activities and ATP amounts in treated animal models.

Synergistic Neuroprotective Effects of Mature Silkworm and Angelica gigas Against Scopolamine-Induced Mild Cognitive Impairment in Mice and H2O2-Induced Cell Death in HT22 Mouse Hippocampal Neuronal Cells.

We previously reported that mature Bombyx mori silkworm (SW) ameliorated scopolamine (Sco)-induced amnesia, and Angelica gigas (AG) prevented cognitive impairment. SW is known for its gastroprotective effects such as improving liver function and alleviating the effects of Parkinson's disease. AG is known for its neuroprotective effects and for lowering the effects of low-density lipoprotein cholesterol. However, the neuroprotective effect of combined SW and AG (SWA-1) treatment and the underlying molecular mechanism by which SWA-1 regulates neurodegenerative diseases remains unclear. We evaluated the neuroprotective effect of SWA-1 against Sco-induced mild cognitive impairment in mice and H2O2-induced cell death in HT22 mouse hippocampal neuronal cells and elucidated the underlying molecular mechanism. Morris water maze and Y-maze tests were performed to examine the learning and memory abilities of mice. The underlying molecular mechanism was investigated by using western blotting. We demonstrated that SWA-1 significantly protects against H2O2-induced cell death in HT22 mouse hippocampal neuronal cells. SWA-1 also significantly reversed Sco-induced spatial learning and memory impairment. Specifically, SWA-1 upregulates the protein levels of phosphorylated extracellular signal-related kinase (Erk1/2) and phosphorylated p38 MAP kinase (p38). SWA-1 remarkably decreased the apoptotic index Bax/Bcl2 expression in the hippocampus of Sco-treated mice. Our results suggest that SWA-1 may be administered as alternative therapy for cognitive impairment and neurodegenerative diseases and should be studied further in human trials.

Effect of ACG-1, an Extract Blend of Angelica gigas, Cynanchum wilfordii, and Ginkgo biloba, on Blood Circulation Improvement Via Antiplatelet Aggregation and Antihyperlipidemia.

Thrombosis causes poor blood circulation, which may lead to several cardiovascular disorders. Antiplatelet aggregation and antihyperlipidemia are the key processes that improve blood circulation. The antiplatelet aggregation and antihyperlipidemic effects of ACG-1, a mixture of Angelica gigas, Cynanchum wilfordii, and Ginkgo biloba extracts, were investigated in this study. The antiplatelet aggregation activity of ACG-1 was determined by studying its effects on collagen-induced platelet aggregation in human platelet-rich plasma (PRP). In addition, the effects of ACG-1 were investigated in a thromboembolism mouse model. The high-fat diet (HFD)-fed mouse model was used to investigate the antihyperlipidemic effects of ACG-1 and western blotting assay was performed to elucidate its mechanism of action. It was observed that ACG-1 significantly inhibited platelet aggregation in human PRP. Furthermore, ACG-1 showed protective effects in a thromboembolism mouse model induced by administering a mixed collagen and epinephrine intravenous injection. Oral administration of ACG-1 also significantly ameliorated blood lipid profiles in the HFD-fed mouse model. In conclusion, ACG-1 should be considered a powerful functional food to improve blood circulation.

The Effect of High Dose Intravenous Vitamin C During Radiotherapy on Breast Cancer Patients' Neutrophil-Lymphocyte Ratio.

Background: Breast cancer is very common, and the incidence is growing every year. Most breast cancers are treated with radiation after surgery. As a side effect of radiation therapy, inflammation, as well as the neutrophil-lymphocyte ratio (NLR), level increases. However, high NLR levels act as independent prognostic factors for increased mortality in all cancers. In this study, the authors investigated whether administration of vitamin C, which is effective in suppressing inflammation, may help to reduce high levels of NLR produced by radiation therapy. Methods: This study was performed retrospectively among 424 patients who were diagnosed with breast cancer and were treated with postoperative radiotherapy at Kosin University Gospel Hospital from January 2011 to December 2017. Among them, 354 patients received radiation therapy without vitamin C (the control group), and 70 experimental patients received vitamin C intravenously twice a week for at least 4 weeks during radiation therapy. The experimental group was divided into two groups according to the dose administrated: a low-dose vitamin C group (less than 1 g/kg, 52 patients) and a high-dose vitamin C group (more than 1 g/kg, 18 patients). The authors conducted three NLR measurements: before and after radiation therapy and at 3 months after radiation therapy; the authors then compared the change in NLR over time between the groups using repeated measures analysis of variance. Results: In the control group and the low-dose vitamin C-administered group, NLR was increased at the endpoint compared to before the radiotherapy, whereas NLR values in the high-dose vitamin C group were 8.4 ± 1.7, 5.9 ± 1.3, and 4.3 ± 1.5, showing a continuous decrease and a statistically significant difference (pinteraction = 0.033). These results were similarly observed in models adjusted by the patient's age and American Joint Committee on Cancer stage, with borderline significance (pinteraction = 0.065). Conclusions: Elevated NLR, a measure of systemic inflammation, has been associated with higher mortality cancer patients, including breast cancer patients. In this observational study, NLR was significantly decreased during radiation therapy in patients administered high-dose vitamin C.

Bee venom inhibits the proliferation and migration of cervical-cancer cells in an HPV E6/E7-dependent manner.

Bee venom (BV), secreted from the venom gland of the honey bee, contains several biological active compounds. BV has been widely used as a traditional medicine for treating human disease, including cancer. In this study, we have shown the molecular mechanism underlying the therapeutic effect of BV on cancer. Treatment with BV reduced the proliferation of cervical-cancer cells in a dose- and time-dependent manner. Interestingly, the killing effect of BV was specific to HPVpositive cervical-cancer cell lines, such as Caski and HeLa cells, and not to HPV-negative cervical-cancer cells (C33A). BV reduced the expression of HPV E6 and E7 at RNA and protein levels, leading to an increase in the expression of p53 and Rb in Caski and HeLa cells. Further, BV decreased the levels of cell-cycle proteins, such as cyclin A and B, and increased the levels of cell-cycle inhibitors, such as p21 and p27. BV significantly induced apoptosis and inhibited wound healing and migration of cervical-cancer cells. It also upregulated the expression of pro-apoptotic BAX and downregulated the expression of anti-apoptotic Bcl-2 and Bcl-XL. Cleavage of caspase-3, caspase-9, and PARP were also induced by BV treatment, whereas the phosphorylation of mitogenic signalingrelated proteins, such as AKT, JNK, p38, and ERK, were downregulated. Our results indicate that BV has a therapeutic selectivity for HPV-positive malignant cells, so further clinical studies are needed to assess its clinical application. [BMB Reports 2020; 53(8): 419-424].

Evaluating the Memory Enhancing Effects of Angelica gigas in Mouse Models of Mild Cognitive Impairments.

(1) Background: By 2050, it is estimated that 130 million people will be diagnosed with dementia, and currently approved medicines only slow the progression. So preventive intervention is important to treat dementia. Mild cognitive impairment is a condition characterized by some deterioration in cognitive function and increased risk of progressing to dementia. Therefore, the treatment of mild cognitive impairment (MCI) is a possible way to prevent dementia. Angelica gigas reduces neuroinflammation, improves circulation, and inhibits cholinesterase, which can be effective in the prevention of Alzheimer's disease and vascular dementia and the progression of mild cognitive impairment. (2) Methods: Angelica gigas (AG) extract 1 mg/kg was administered to mildly cognitive impaired mice, models based on mild traumatic brain injury and chronic mild stress. Then, spatial, working, and object recognition and fear memory were measured. (3) Result: Angelica gigas improved spatial learning, working memory, and suppressed fear memory in the mild traumatic brain injury model. It also improved spatial learning and suppressed cued fear memory in the chronic mild stress model animals. (4) Conclusions: Angelica gigas can improve cognitive symptoms in mild cognitive impairment model mice.

Expression of the Arabidopsis WRINKLED 1 transcription factor leads to higher accumulation of palmitate in soybean seed.

Soybean (Glycine max [L.] Merr.) is a commodity crop highly valued for its protein and oil content. The high percentage of polyunsaturated fatty acids in soybean oil results in low oxidative stability, which is a key parameter for usage in baking, high temperature frying applications, and affects shelf life of packaged products containing soybean oil. Introduction of a seed-specific expression cassette carrying the Arabidopsis transcription factor WRINKLED1 (AtWRI1) into soybean, led to seed oil with levels of palmitate up to approximately 20%. Stacking of the AtWRI1 transgenic allele with a transgenic locus harbouring the mangosteen steroyl-ACP thioesterase (GmFatA) resulted in oil with total saturates up to 30%. The creation of a triple stack in soybean, wherein the AtWRI1 and GmFatA alleles were combined with a FAD2-1 silencing allele led to the synthesis of an oil with 28% saturates and approximately 60% oleate. Constructs were then assembled that carry a dual FAD2-1 silencing element/GmFatA expression cassette, alone or combined with an AtWRI1 cassette. These plasmids are designated pPTN1289 and pPTN1301, respectively. Transgenic events carrying the T-DNA of pPTN1289 displayed an oil with stearate levels between 18% and 25%, and oleate in the upper 60%, with reduced palmitate (<5%). While soybean events harboring transgenic alleles of pPTN1301 had similar levels of stearic and oleate levels as that of the pPTRN1289 events, but with levels of palmitate closer to wild type. The modified fatty acid composition results in an oil with higher oxidative stability, and functionality attributes for end use in baking applications.

Woohwangcheongsimwon Prevents High-Fat Diet-Induced Memory Deficits and Induces SIRT1 in Mice.

Woohwangcheongsimwon (WHC) is a mixture of herbal medicines that is widely prescribed in Korean traditional medicine. SIRT1 is known for its regulatory roles in energy metabolism, oxidative stress, and circadian rhythms. This study was designed to determine whether WHC can increase and mimic the biological reactions of SIRT1 activation. Ten-month-old male mice were divided into four groups: nontreated normal diet (ND), nontreated high-fat diet (HFD), WHC-treated ND, and WHC-treated HFD. Body weight and cognitive functions were evaluated after treatment. The hippocampal expressions of SIRT1 and PGC-1α were also measured. The components of WHC were identified by liquid chromatography. High-fat diet-fed mice gained more weight and demonstrated greater deficits in short-term and long-term cognitive functions. WHC suppressed the deleterious effects of a HFD on weight gain and cognitive decline, but showed no prominent effects on animals fed NDs. The herbal treatment also increased the expression of SIRT1 and PGC-1α in the hippocampus. Despite the induction of hippocampal SIRT1 expression by WHC, resveratrol was not present among the natural compounds identified. This expression might have contributed to the suppression of high-fat diet-induced memory deficits in mice treated with the herbal mixture.

Towards the development of a sustainable soya bean-based feedstock for aquaculture.

Soya bean (Glycine max (L.) Merr.) is sought after for both its oil and protein components. Genetic approaches to add value to either component are ongoing efforts in soya bean breeding and molecular biology programmes. The former is the primary vegetable oil consumed in the world. Hence, its primary usage is in direct human consumption. As a means to increase its utility in feed applications, thereby expanding the market of soya bean coproducts, we investigated the simultaneous displacement of marine ingredients in aquafeeds with soya bean-based protein and a high Omega-3 fatty acid soya bean oil, enriched with alpha-linolenic and stearidonic acids, in both steelhead trout (Oncorhynchus mykiss) and Kampachi (Seriola rivoliana). Communicated herein are aquafeed formulations with major reduction in marine ingredients that translates to more total Omega-3 fatty acids in harvested flesh. Building off of these findings, subsequent efforts were directed towards a genetic strategy that would translate to a prototype design of an optimal identity-preserved soya bean-based feedstock for aquaculture, whereby a multigene stack approach for the targeted synthesis of two value-added output traits, eicosapentaenoic acid and the ketocarotenoid, astaxanthin, were introduced into the crop. To this end, the systematic introduction of seven transgenic cassettes into soya bean, and the molecular and phenotypic evaluation of the derived novel events are described.

Metabolic engineering of oilseed crops to produce high levels of novel acetyl glyceride oils with reduced viscosity, freezing point and calorific value.

Seed oils have proved recalcitrant to modification for the production of industrially useful lipids. Here, we demonstrate the successful metabolic engineering and subsequent field production of an oilseed crop with the highest accumulation of unusual oil achieved so far in transgenic plants. Previously, expression of the Euonymus alatus diacylglycerol acetyltransferase (EaDAcT) gene in wild-type Arabidopsis seeds resulted in the accumulation of 45 mol% of unusual 3-acetyl-1,2-diacyl-sn-glycerols (acetyl-TAGs) in the seed oil (Durrett et al., 2010 PNAS 107:9464). Expression of EaDAcT in dgat1 mutants compromised in their ability to synthesize regular triacylglycerols increased acetyl-TAGs to 65 mol%. Camelina and soybean transformed with the EaDAcT gene accumulate acetyl-triacylglycerols (acetyl-TAGs) at up to 70 mol% of seed oil. A similar strategy of coexpression of EaDAcT together with RNAi suppression of DGAT1 increased acetyl-TAG levels to up to 85 mol% in field-grown transgenic Camelina. Additionally, total moles of triacylglycerol (TAG) per seed increased 20%. Analysis of the acetyl-TAG fraction revealed a twofold reduction in very long chain fatty acids (VLCFA), consistent with their displacement from the sn-3 position by acetate. Seed germination remained high, and seedlings were able to metabolize the stored acetyl-TAGs as rapidly as regular triacylglycerols. Viscosity, freezing point and caloric content of the Camelina acetyl-TAG oils were reduced, enabling use of this oil in several nonfood and food applications.

Redirection of metabolic flux for high levels of omega-7 monounsaturated fatty acid accumulation in camelina seeds.

Seed oils enriched in omega-7 monounsaturated fatty acids, including palmitoleic acid (16:1∆9) and cis-vaccenic acid (18:1∆11), have nutraceutical and industrial value for polyethylene production and biofuels. Existing oilseed crops accumulate only small amounts (<2%) of these novel fatty acids in their seed oils. We demonstrate a strategy for enhanced production of omega-7 monounsaturated fatty acids in camelina (Camelina sativa) and soybean (Glycine max) that is dependent on redirection of metabolic flux from the typical ∆9 desaturation of stearoyl (18:0)-acyl carrier protein (ACP) to ∆9 desaturation of palmitoyl (16:0)-acyl carrier protein (ACP) and coenzyme A (CoA). This was achieved by seed-specific co-expression of a mutant ∆9-acyl-ACP and an acyl-CoA desaturase with high specificity for 16:0-ACP and CoA substrates, respectively. This strategy was most effective in camelina where seed oils with ~17% omega-7 monounsaturated fatty acids were obtained. Further increases in omega-7 fatty acid accumulation to 60-65% of the total fatty acids in camelina seeds were achieved by inclusion of seed-specific suppression of 3-keto-acyl-ACP synthase II and the FatB 16:0-ACP thioesterase genes to increase substrate pool sizes of 16:0-ACP for the ∆9-acyl-ACP desaturase and by blocking C18 fatty acid elongation. Seeds from these lines also had total saturated fatty acids reduced to ~5% of the seed oil versus ~12% in seeds of nontransformed plants. Consistent with accumulation of triacylglycerol species with shorter fatty acid chain lengths and increased monounsaturation, seed oils from engineered lines had marked shifts in thermotropic properties that may be of value for biofuel applications.

Stacking of a stearoyl-ACP thioesterase with a dual-silenced palmitoyl-ACP thioesterase and ∆12 fatty acid desaturase in transgenic soybean.

Soybean (Glycine max (L.) Merr) is valued for both its protein and oil, whose seed is composed of 40% and 20% of each component, respectively. Given its high percentage of polyunsaturated fatty acids, linoleic acid and linolenic acid, soybean oil oxidative stability is relatively poor. Historically food processors have employed a partial hydrogenation process to soybean oil as a means to improve both the oxidative stability and functionality in end-use applications. However, the hydrogenation process leads to the formation of trans-fats, which are associated with negative cardiovascular health. As a means to circumvent the need for the hydrogenation process, genetic approaches are being pursued to improve oil quality in oilseeds. In this regard, we report here on the introduction of the mangosteen (Garcinia mangostana) stearoyl-ACP thioesterase into soybean and the subsequent stacking with an event that is dual-silenced in palmitoyl-ACP thioesterase and ∆12 fatty acid desaturase expression in a seed-specific fashion. Phenotypic analyses on transgenic soybean expressing the mangosteen stearoyl-ACP thioesterase revealed increases in seed stearic acid levels up to 17%. The subsequent stacked with a soybean event silenced in both palmitoyl-ACP thioesterase and ∆12 fatty acid desaturase activity, resulted in a seed lipid phenotype of approximately 11%-19% stearate and approximately 70% oleate. The oil profile created by the stack was maintained for four generations under greenhouse conditions and a fifth generation under a field environment. However, in generation six and seven under field conditions, the oleate levels decreased to 30%-40%, while the stearic level remained elevated.

Preclinical evidence of rapid-onset antidepressant-like effect in Radix Polygalae extract.

Radix Polygalae (the root of Polygala tenuifolia) is a herb widely used in traditional Asian medicine that is thought to exert a variety of neuropsychiatric effects. Radix Polygalae extract can protect against N-methyl D-aspartate (NMDA) neurotoxicity and induce brain-derived neurotrophic factor (BDNF) expression, suggesting modulatory roles at glutamatergic synapses and possible antidepressant action. In accordance with this hypothesis, Radix Polygalae extract demonstrated antidepressant-like effects in 8-week-old male C57Bl/6 mice by decreasing behavioral despair in the forced swim and tail suspension tasks and increasing hedonic-like behavior in the female urine sniffing test 30 minutes after a single oral administration of 0.1 mg/kg. Reduced latency to acquire a food pellet in the novely suppressed feeding paradigm, without change in anxiety-like behaviors suggested a rapid-onset nature of the antidepressant-like effect. In addition, it decreased the number of failed escapes in the learned helplessness paradigm after two oral administrations 24 hours and 30 minutes before the first test. Finally, it reversed anhedonia as measured by saccharin preference in mice exposed to the chronic stress model after two administrations of 0.1 mg/kg, in contrast to the repeated administration generally needed for similar effect by monoamergic antidepressants. Immobility reduction in tail suspension task was blocked by the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor antagonist NBQX, a pattern previously demonstrated by ketamine and other ketamine-like rapid-onset antidepressants. Also similarly to ketamine, Radix Polygalae appeared to acutely decrease phosphorylation of GluR1 serine-845 in the hippocampus while leaving the phosphorylation of hippocampal mTOR serine 2448 unchanged. These findings serve as preclinical evidence that Radix Polygalae extract exerts rapid-onset antidepressant effects by modulating glutamatergic synapses in critical brain circuits of depression and may be worthy of further evaluation as a safe substitute to other rapid-onset antidepressants known to have unacceptable side effects.

The complete chloroplast genome sequences of Solanum tuberosum and comparative analysis with Solanaceae species identified the presence of a 241-bp deletion in cultivated potato chloroplast DNA sequence.

The complete nucleotide sequence of the chloroplast genome of potato Solanum tuberosum L. cv. Desiree was determined. The circular double-stranded DNA, which consists of 155,312 bp, contains a pair of inverted repeat regions (IRa, IRb) of 25,595 bp each. The inverted repeat regions are separated by small and large single copy regions of 18,373 and 85,749 bp, respectively. The genome contains 79 proteins, 30 tRNAs, 4 rRNAs, and unidentified genes. A comparison of chloroplast genomes of seven Solanaceae species revealed that the gene content and their relative positions of S. tuberosum are similar to the other six Solanaceae species. However, undefined open reading frames (ORFs) in LSC region were highly diverged in Solanaceae species except N. sylvestris. Detailed comparison was identified by numerous indels in the intergenic regions that were mostly located in the LSC region. Among them, a single large 241-bp deletion, was not associated with direct repeats and found in only S. tuberosum, clearly discriminates a cultivated potato from wild potato species Solanum bulbocastanum. The extent of sequence divergence may provide the basis for evaluating genetic diversity within the Solanaceae species, and will be useful to examine the evolutionary processes in potato landraces.

Effect of genistein with carnitine administration on lipid parameters and obesity in C57Bl/6J mice fed a high-fat diet.

Soy products are mainly composed of proteins, phytochemicals such as isoflavones, soy lipids, and carbohydrates. It is unclear whether an individual component alone or a combined effect of multiple bioactive compounds contributes to the beneficial properties of soy. We investigated the effect of dietary genistein (the principal soy isoflavone) alone and combined with L-carnitine to evaluate possible synergistic effects on the intentionally induced prediabetic state characterized by insulin resistance and obesity in C57Bl/6J mice fed a high-fat diet (HD). In the HD-alone group, abdominal and back fat relative to total body weight were significantly higher compared with other groups including those fed normal diet (ND). Among the HD groups, final weight gains of the HD plus genistein (HD+G) and HD plus genistein plus L-carnitine (HD+G+C) groups were lower compared with that of the control (HD-alone). Especially in liver, the results showed that genistein with carnitine transcriptionally up-regulated expressions of acyl-coenzyme A synthetase (ACS) and carnitine palmitoyltransferase-I (CPT-I) by approximately 50% and 40%, respectively, compared with genistein alone. However, the up-regulation of CPT-I did not directly reflect the enzyme activity of CPT-I. On the other hand, the effects of genistein and genistein with carnitine on the expressions of ACS and CPT-I in muscle were not significant. Our study suggests that genistein with carnitine exerts anti-obesity effects, probably by modulating peroxisome proliferator-activated receptor-associated genes. However, further work is needed to elucidate the possible mechanisms by which genistein and carnitine intervene.

Analysis of transcripts in methyl jasmonate-treated ginseng hairy roots to identify genes involved in the biosynthesis of ginsenosides and other secondary metabolites.

Methyl jasmonate (MeJA) treatment increases the levels of plant secondary metabolites, including ginsenosides, which are considered to be the main active compounds in ginseng (Panax ginseng C.A. Meyer). To create a ginseng gene resource that contains the genes involved in the biosynthesis of secondary metabolites, including ginsenosides, we generated 3,134 expression sequence tags (ESTs) from MeJA-treated ginseng hairy roots. These ESTs assembled into 370 clusters and 1,680 singletons. Genes yielding highly abundant transcripts were those encoding proteins involved in fatty acid desaturation, the defense response, and the biosynthesis of secondary metabolites. Analysis of the latter group revealed a number of genes that may be involved in the biosynthesis of ginsenosides, namely, oxidosqualene cyclase (OSC), cytochrome P450, and glycosyltransferase. A novel OSC gene was also identified by this analysis. RNA gel blot analysis confirmed that transcription of this OSC gene, along with squalene synthase (SS) and squalene epoxidase (SE) gene transcription, is increased by MeJA treatment. This ginseng EST data set will also provide important information on the genes that are involved in the biosynthesis of other secondary metabolites and the genes that are responsive to MeJA treatment.