Identification, Characterization and Expression Analysis of Anthocyanin Biosynthesis-related bHLH Genes in Blueberry (Vaccinium corymbosum L.)

Basic helix-loop-helix proteins (bHLHs) play very important roles in the anthocyanin biosynthesis of many plant species. However, the reports on blueberry anthocyanin biosynthesis-related bHLHs were very limited. In this study, six anthocyanin biosynthesis-related bHLHs were identified from blueberry genome data through homologous protein sequence alignment. Among these blueberry bHLHs, VcAN1, VcbHLH42-1, VcbHLH42-2 and VcbHLH42-3 were clustered into one group, while VcbHLH1-1 and VcbHLH1-2 were clustered into the other group. All these bHLHs were of the bHLH-MYC_N domain, had DNA binding sites and reported conserved amino acids in the bHLH domain, indicating that they were all G-box binding proteins. Protein subcellular location prediction result revealed that all these bHLHs were nucleus-located. Gene structure analysis showed that VcAN1 gDNA contained eight introns, while all the others contained seven introns. Many light-, phytohormone-, stress- and plant growth and development-related cis-acting elements and transcription factor binding sites (TFBSs) were identified in their promoters, but the types and numbers of cis-elements and TFBSs varied greatly between the two bHLH groups. Quantitative real-time PCR results showed that VcAN1 expressed highly in old leaf, stem and blue fruit, and its expression increased as the blueberry fruit ripened. Its expression in purple podetium and old leaf was respectively significantly higher than in green podetium and young leaf, indicating that VcAN1 plays roles in anthocyanin biosynthesis regulation not only in fruit but also in podetium and leaf. VcbHLH1-1 expressed the highest in young leaf and stem, and the lowest in green fruit. The expression of VcbHLH1-1 also increased as the fruit ripened, and its expression in blue fruit was significantly higher than in green fruit. VcbHLH1-2 showed high expression in stem but low expression in fruit, especially in red fruit. Our study indicated that the anthocyanin biosynthesis regulatory functions of these bHLHs showed certain spatiotemporal specificity. Additionally, VcAN1 might be a key gene controlling the anthocyanin biosynthesis in blueberry, whose function is worth exploring further for its potential applications in plant high anthocyanin breeding.

Differences Between Healthy and Ganoderma boninense Infected Oil Palm Seedlings Using Spectral Reflectance of Young Leaf Data

Ganoderma boninense (G.boninense) is the causal agent of basal stem rot (BSR) which significantly reduced the productivity of oil palm plantations in Southeast Asia. At early stage, the disease did not show any physical symptoms that could be seen with naked eyes resulted in detection difficulties. To date, there was no effective detection for this disease, and conventional methods such as manual and laboratory-based required trained specialists as well as time-consuming. Therefore, this study was conducted using hyperspectral remote sensing to investigate the differences in spectral reflectance of young leaf (frond one (F1) of healthy and G. boninense infected oil palm seedlings. The seedlings were inoculated with G. boninense pathogen at five months old. At five months after inoculation, 558 spectral signatures of F1 were extracted from acquired hyperspectral images. Noise removal was done to the extracted spectral signatures to remove outliers in the data. Then, the spectral signatures were averaged and plotted to observe the differences. Differences in reflectance of healthy and G. boninense infected seedlings were seen evidently in the near-infrared (NIR) region. Thus, this study showed evidence that F1 spectral reflectance has the ability to detect early stage of G. boninense infection at oil palm seedlings.

Comprehensive Transcriptome and Metabolome Analysis of Lithocarpus polystachyus Leaf Revealed Key Genes in Flavonoid Biosynthesis Pathways

Lithocarpus polystachyus is a unique medicinal tree species that is valued for its abundant flavonoids in leaves. Currently, genes and metabolites involved in the flavonoid biosynthesis pathway remain largely unknown. To elucidate the flavonoid biosynthesis pathways, transcriptome and metabolome analyses of young, mature, and old leaves were conducted. A total of 86,927 unigenes were obtained, and 51.4% of them were annotated in eight public databases. The majority of the 44 candidate genes in the flavonoid biosynthesis pathway were downregulated as leaves aged. Metabolome profiling revealed a set of 427 metabolites in leaves. Consistent with the transcriptome results, 15 of the 19 metabolites in the flavonoid pathway decreased during the development of leaves. The data indicate that young leaf is the optimal stage for tea harvest. This is the first report of integrated transcriptome and metabolome profiling of L. polystachyus. This study demonstrates the correlation of gene expression and metabolites related to flavonoid biosynthesis and reveals the key genes responsible for flavonoid accumulation in young leaf. The information can be applied to future studies performed to elucidate and manipulate flavonoid biosynthesis in L. polystachyus.

Establishment of plant regeneration system for young leaf explants of Cassia mimosoides

In the present study, a plant regeneration system was established by using young leaf explants from aseptic seedlings of Cassia mimosoides. The results showed that when the explants were inoculated on the 1/2MS medium contained 2.0 mg/L 6-BA and 0.5 mg/L NAA, high rate of callus induction and good growth status of adventitious bud, and the highest number of buds were obtained. Further studies conclusively suggested that IBA could not promote root formation effectively, but the root regeneration could be induced by NAA. However, although the rooting of regeneration buds could be induced when adventitious buds were placed into 1/2MS medium supplemented with 0.5 mg/L NAA, the rate of rooting was reduced, and the roots became shorter. Therefore, the best medium for adventitious bud regeneration using young leaf as explants in C. mimosoides was 1/2MS medium contained 2.0 mg/L 6-BA and 0.5 mg/L NAA, and the best rooting medium was 1/2MS.

Kadar Mangiferin Pada Lima Kultivar Pucuk Daun Mangga (Mangifera indica L.)

AbstrakPemanfaatan pucuk daun mangga (Mangifera indica L.) oleh masyarakat menjadi lalapan dan dipercaya berkhasiat sebagai obat tradisional. Hal ini merupakan suatu proses kearifan lokal berdasarkan pengalaman empiris orang-orang terdahulu yang kemudian dijadikan kajian studi ilmiah, yaitu etnobotani. Kajian studi etnobotani termasuk pemanfaatan pucuk daun mangga sebagai lalapan masih jarang dilakukan. Penelitian ini bertujuan untuk mengetahui senyawa bioaktif termasuk mangiferin yang terkandung dalam lima kultivar mangga. Sampel kultivar mangga yang digunakan dalam penelitian antara lain Manalagi, Arum manis, Cengkir, Gedong apel dan Golek. Metode penelitian yang digunakan yaitu skrining fitokimia dan pengukuran kadar senyawa mangiferin dengan HPLC. Hasil penelitian menunjukkan sampel positif mengandung senyawa bioaktif alkaloid, flavonoid, saponin dan tanin. Kadar senyawa mangiferin paling tinggi secara berurutan terdapat pada kultivar Arum manis (20,83%), Manalagi (20,56%), Gedong apel (18,54%), Cengkir (15,10%), dan Golek (5,05%). Kultivar mangga Arum manis memiliki kadar mangiferin paling tinggi, untuk selanjutnya dapat dikaji pemanfaatannya sebagai obat herbal tradisional. Abstract Utilization young leaf of mango (Mangifera indica L.) by the community as lalapan and it is believed to be efficacious as traditional medicines. This process through of local wisdom based on empirical experience of conservative people used as a scientific study, namely ethnobotany. Ethnobotany studies include the use of mango leaf shoots as fresh vegetables are still rarely performed.This study aims to determine bioactive compounds including mangiferin contained in five cultivars of mango. Mango cultivar samples used in this study include Manalagi, Arum manis, Cengkir, Gedong apel and Golek. The research method used was phytochemical screening and measurement of mangiferin compounds by HPLC. The results showed a positive sample containing bioactive compounds, alkaloids, flavonoids, saponins and tannins. The highest levels of mangiferin compounds were found in Arum manis (20.83%), Manalagi (20.56%), Gedong apel (18.54%), Cengkir (15.10%) and Golek (5.05%). Arum manis cultivars has highest value of mangiferin levels, which can be explored for their use as traditional herbal medicines.

CitRWP May not be the Major or Only Major Gene that Regulates Apomixis in Citrus and Its Close Related Genus

Background: Nucellar embryony in citrus and its close related genus is a kind of apomixis, it interferes the cross breeding and affects the genetic evolution and taxonomy in citrus. Many hypotheses have been suggested on regulation of apomixes in citrus but none of them are able to lock the true regulatory sequence until the recently reported candidate gene citRWP. However, the support evidences for the judgment of citRWP as the major or only major gene are still not very sound.Results: Firstly, M/P (Monoembryonic / Polyembryonic) marker, which was designed according to the upper stream sequence of citRWP, was used to check whether the marker exhibiting consistent segregation in different embryonic genotypes. The result showed that all sweet orange selections either monoembryonic or polyembryonic contained the P-marker, but no P-marker was detected in polyembryonic Poncirus, and a new band pattern was dectected in 9 polyembryonic progenies of W-Murcott × Flying dragon trifoliate orange. Subsequently, CitRWP expressions were analyzed with RT-PCR and Realtime RT-PCR. RT-PCR revealed that CitRWP expressed in ovule of all varieties. Realtime RT-PCR results in ovule were consistent with RT-PCR. The expression value of monoembryonic pummelo was the lowest, and there was no significant difference between polyembroynic and monoembryonic sweet orange cultivars. In the hybrid offsprings, the expression of the CitRWP gene showed an extreme two-stage differentiation. The expression in young leaf and ovule of polyembryonic and monoembryonic sweet orange cultivars were different, with a higher expression level in young leaf and a lower expression level in ovule of polyembryonic cultivars. Finally, the cDNA sequences of CitRWP from ovules were cloned and sequenced. Results revealed that the CitRWP has great variations in Poncirus, which included SNPs in start-codon and/or stop-codon as well as fragment insertion/deletion causing incorrect translation. The monoembryonic/polyembryonic sweet orange cultivars and pummelo varieties had similar variations also.Conclusion: CitRWP were not consistently distributed and expressed in different embryonic genotypes. It may not be the major or only major gene that regulates apomixis in citrus and its close related genus.

PERKEMBANGAN BIBIT AREN (Arenga pinnata Merr) YANG DIKULTUR PADA MEDIA MS DAN WPM

Arenga palm (Arenga pinnata) is one species from palmae family. Economically, the palm is suitable for home industry, such as Balinese offering which use parts of young leaf, sap can be used as special local genius sweet drink from Bali, fruits are boilled that can be mixed with sugar for healthy ice. Arenga palm can be propagated aseptically using tissue culture technique. Culture media may varried between MS and WPM with or without adding hormone. Auxin can be added as 2.4-D ( 4 ppm) to induce callus. NAA and BAP with concentration 1.5 ppm respectively also added to induce multiplication of shoots or roots. Each treatment had 10 replicatations. Six weeks after cultured, callus growth observed only at WPM media that enriched with 2,4-D. The others treatment show the explant was dormant because did not show any growth while analysed anatomically under microscope.

Antimicrobial Activity of Methanolic, Aqueous and Partially Purified Protein of Young and Matured Leaves of Guiera senegalensis (Moshi Medicine)

Background: Microorganisms have evolved defence mechanisms against antimicrobial agents and have become resistant to some antibiotics. Aim of the Study: This study aimed to evaluate the antimicrobial activity of methanol as well as that of the aqueous and partially purified protein of young and matured leaves of Guiera senegalensis. Study Duration: This study was conducted on 30th January, 2015 at the Department of Biochemistry, Faculty of Science, Ahmadu Bello University Zaria, Nigeria. Methodology: Antimicrobial activity was determined using disc diffusion and broth dilution techniques, Gel chromatography techniques were used to fractionate the crude protein. Results: The Test isolates were Bacillus subtilis, Staphylococcus aureus, E. coli, Salmonella typhimurium and Candida albicans. All the extracts were sensitive to most of the isolates except Candida albican (fungus). The crude and partially purified proteins were active against the Gram positive bacteria. The maximum zone of inhibition (42.00±1.00 mm) was observed in the methanol extract of a young leaf against Staphylococcus aureus at 100 mg/ml. The methanol extracts exhibited minimum inhibitory concentration (MIC) at a range of 6.25 mg/ml and 12.5 mg/ml and minimum bactericidal concentration (MBC) at 12.5 mg/ml and 25 mg/ml. The young leaf was more active than the matured leaf. Quantitative phytochemicals showed high amounts of saponins (26.20% and 19.66%) in matured and young leaves respectively. Conclusion: This research justifies the traditional claim of Guiera senegalensis leaves for therapeutic purposes. The leaves can be used to remedy diseases caused by bacterial agents.