Sony Suhandono

PCR primers amplifying microsatellite loci of Quercus myrsinifolia Blume and their conservation between oak species

Approximately 450 Quercus species are known around the world. Most of them are important members of forest ecosystems that determine the structure of the mixed mountain forests of the world. Because of their economic and ecological importance, much research on the regeneration process has been carried out. In addition, Quercus species are known to conduct interesting reproductive behaviour, masting: synchronous large seed production at long intervals by a population of plants (Kelly 1994), and the evolutionary benefits of this behaviour have been proposed (Janzen 1971; Waller 1979; Silvertown 1980; Nilsson & Watljung 1987; Norton & Kelly 1988; Ims 1990; Koenig et a/ . 1994). In order to evaluate the ecological and evolutionary benefits of the masting behaviour, we have to determine the difference in gene flow between parents of masting or nonmasting individuals to seedlings. Microsatellites are short, 1-6 bp in length, tandemly repeated simple sequence DNA (Ashley & Dow 1994), and can be applied to ecological and evolutionary studies because they are highly variable, codominantly inherited genetic markers segregating at distinct loci. For cultivated plants, a sufficient number of microsatellite markers has been developed. However, such markers have been relatively rare for wild plants. Quercus rnyrsinifolia is an evergreen oak, and constructs a canopy of evergreen forests in Japan and eastern China. Like most Quercus species, Q. rnyrsinifolia produces an intermittent large seed crop. For Quercus species, three sets of primers were recently reported (Dow et a/ . 1995). The number of sets is, however, insufficient to conduct an analysis of the genetic structure of a wild population. We report here a sufficient number of primer sets amplifying microsatellite sites for the paternity and population analysis of Qzwrms rnyrsirrifolia.

Measuring leaf chlorophyll concentration from its color: A way in monitoring environment change to plantations

Leaf colors of a plant can be used to identify stress level due to its adaptation to environmental change. For most leaves green-related colors are sourced from chlorophyll a and b. Chlorophyll concentration is normally measured using a spectrophotometer in laboratory. In some remote observation places, it is impossible to collect the leaves, preserve them, and bring them to laboratory to measure their chlorophyll content. Based on this need, measurement of chlorophyll content is observed through its color. Using CIE chromaticity diagram leaf_color information in RGB is transformed into wavelength (in nm). Paddy seed with variety name IR-64 is used in observation during its vegetation stage t (age of 0-10 days). Light exposure time τ is chosen as environmental change, which normally should be about 12 hours/day, is varied (0-12 hours/day). Each day sample from different exposure time is taken, its color is recorded using HP Deskjet 1050 scanner with 1200 dpi, and its chlorophyll content is obtained from a...

Optimization of genetic transformation of Artemisia annua L. Using Agrobacterium for Artemisinin production.

Background: Artemisinin, a sesquiterpene lactone endoperoxide isolated from the medicinal plant Artemisia annua L., is a choice and effective drug for malaria treatment. Due to the low yield of artemisinin in plants, there is a need to enhance the production of artemisinin from A. annua and biotechnological technique may be one of the methods that can be used for the purpose. Aim: To study the transformation efficiency of Agrobacterium tumefaciens in A. annua that could be applied to enhance the production of artemisinin by means of transgenic plants. Setting and Designs: The factors influencing Agrobacterium-mediated transformation of A. annua were explored to optimize the transformation system, which included A. tumefaciens strain and effect of organosilicone surfactants. Three strains of A. tumefaciens, that is, LBA4404, GV1301, and AGL1 harboring the binary vector pCAMBIA 1303 have been used for transformation. The evaluation was based on transient β-glucuronidase (GUS). Materials and Methods: Plant cell cultures were inniatiated from the seeds of A. annua using the germination Murashige and Skoog medium. A. tumefaciens harboring pCAMBIA were tranformed into the leaves of A.annua cultures from 2-week-old-seedling and 2-month-old-seedling for 15 min by vacuum infiltration. Transformation efficiency was determinated by measuring of blue area (GUS expression) on the whole leaves explant using ImageJ 1.43 software. Two organosilicon surfactants, that is, Silwet L-77 and Silwet S-408 were used to improve the transformation efficiency. Results: The transformation frequency with AGL1 strain was higher than GV3101 and LBA4404 which were 70.91, 49.25, and 45.45%, respectively. Effect of organosilicone surfactants, that is, Silwet L-77 and Silwet S-408 were tested on A. tumefaciens AGL1 and GV3101 for their level of transient expression, and on A. rhizogenes R1000 for its hairy root induction frequency. For AGL1, Silwet S-408 produced higher level of expression than Silwet L-77, were 2.3- and 1.3-fold, respectively. For GV3101, Silwet L-77 was still higher than Silwet S-408, were 1.5- and 1.4-fold, respectively. However, GV3101 produced higher levels of expression than AGL1. The area of GUS expression spots of AGL1, LBA4404, and GV3101 strains was 53.43%, 41.06%, and 30.51%, respectively. Conclusion: A. tumefaciens AGl1 strain was the most effective to be transformed in to A. annua than GV3101 and LBA4404 strain. Surfactant Silwet S-408 produced the highest efficiency of transformation.

Growth Promotion of Rice Plant by Endophytic Fungi

The main aim of this research is to obtain endophytic fungi from graminiae which has the ability to promote the growth of rice plant. The research was conducted in laboratory of Mycology Research Center for BioSciences and Biotechnology Institut Teknologi Bandung. Isolation of endophytic fungi was done by direct plating of root samples on the potatoe dextrose agar media after surface sterilization steps. Identification was based on morphology characteristics and DNA marker. At least 124 isolates were obtained from elephant grass (Pennisetum purpureum) (27 isolates), alang-alang (Imperata cylindrica) (25 isolates), sugar cane (Saccharum officinarum) (14 isolates), maize (Zea mays) (9 isolates), rice plant (Oryza sativa) (23 isolates) and cyperus (Cyperus rotundus) (26 isolates). Among them, 12 isolates demosntrated the capability to promote the growth of rice seedlings in term of seed germination, plant height, root length and degree of root colonization. Isolate I-11 identified as Phialemonium dimorphosphorum was the best to promote rice plant growth in glass house trial.

Bioinformatics Resources in Facilitating Students' Conception on Molecule genetics concepts

Molecule genetics concepts are dificcult concepts for students almost in every country. Students must strugle to visualize those concepts. In this study, we presented bioinformatics resources in facilitating students’ conception on molecule genetics concepts. Our research questions targeted students conception of deoxyriboculeic acid (DNA), transcription, genetic code, codon/anti-codon, intron, exon, gene. Data sources included pre/post-test and transcript of interviews. Our findings on pre-test showed that students have many variatian of molecule genetics concepts as prior knowledge. We found that most students held misconception of all over concepts and have partial conception on intron, exon and gene. Through bioinformatics learning process, the students prior conceptual frameworks was transformed to better understanding. Most students have a comprehensive knowledge of DNA, transcription, genetic code and codon/anti-codon. However most of students still held the partial knowledge of intron, exon and gene. Meanwhile in intron, exon and gene concepts, the students were unable to integrate their prior knowledge with informations of the bioinformatics resources. The transcripts revealed that bioinformatics resources were able to enhance students in representing of the DNA, transcription, genetic code and codon/anti-codon concepts. Keywords—Molecule Genetics; Misconception; Bioinformatics; Learning Process.

Isolation, identification and characterization of indigenous fungi for bioremediation of hexavalent chromium, nickel and cobalt

Waste from nickel mining of Sorowako in South Sulawesi contains hexavalent chromium, nickel and cobalt metals in high concentration and may have a negative impact to the environment. Common waste treatment systems such as chemical treatment using a reducing reagent may still have a negative impact. Bioremediation using fungi or bacteria becomes more popular because it is an environmentally friendly alternative. The purposes of this study are to isolate and identify indigenous fungi that are resistant to heavy metals (hexavalent chromium, nickel, and cobalt) and are capable of reducing the concentration of metals in mining wastes. Ten fungal isolates were successfully isolated from the soils and pond sediments in the area of nickel mining in Sorowako. Selection of superior isolate was carried out by growing all the isolates on PDA medium, which contained all of the three metals. One superior isolate was identified to be able to grow on medium with concentrations of 6400 ppm hexavalent chromium, 200 ppm nickel and 50 ppm cobalt. Molecular identification and phylogenetic studies of the isolate using fungal PCR primers developed to amplify the ITS (internal transcribed spacer) region showed that the isolate sequence was very close to Trichoderma atroviride with 99.8% similarity. Optimum incubation time for the uptake of hexavalent chromium was 3 days, nickel and cobalt was 5 days, respectively, with an optimum pH of 4.Waste from nickel mining of Sorowako in South Sulawesi contains hexavalent chromium, nickel and cobalt metals in high concentration and may have a negative impact to the environment. Common waste treatment systems such as chemical treatment using a reducing reagent may still have a negative impact. Bioremediation using fungi or bacteria becomes more popular because it is an environmentally friendly alternative. The purposes of this study are to isolate and identify indigenous fungi that are resistant to heavy metals (hexavalent chromium, nickel, and cobalt) and are capable of reducing the concentration of metals in mining wastes. Ten fungal isolates were successfully isolated from the soils and pond sediments in the area of nickel mining in Sorowako. Selection of superior isolate was carried out by growing all the isolates on PDA medium, which contained all of the three metals. One superior isolate was identified to be able to grow on medium with concentrations of 6400 ppm hexavalent chromium, 200 ppm nic...

In Planta Transformation Method for T-DNA transfer in Orchids

Transgenic plant technology is an efficient tool to study the function of gene(s) in plant. The most popular and widely used technique is Agrobacterium-mediated transformation in which cocultivation was done by immersing the plant tissues/organ in overnight bacterial cultured for about 30 minutes to one hour under in vitro condition. In this experiment, we developed more easier technique that omitted the in vitro step during cocultivation with Agrobacterium, namely in planta transformation method. Pollinaria (compact pollen mass of orchid) of Phalaenopsis amabilis and Spathoglottis plicata orchids were used as target explants that were immersed into bacterial culture for 30 minutes, then dried up the pollinaria, the transformed pollinaria was used to pollinate orchid flowers. The T-DNA used for this experiments were Ubipro∷PaFT/A. tumefaciens GV3101 for P. amabilis and MeEF1α2 pro∷GUS/ A. tumefaciens LBA 4404 for S.plicata. Seeds that were produced from pollinated flowers were grown onto 10 mg/l hygromici...

Cloning and construction of recombinant palI gene from Klebsiella oxytoca on pET-32b into E. coli BL21 (DE3) pLysS for production of isomaltulose, a new generation of sugar

Klebsiella oxytoca produces sucrose isomerase which catalyses the conversion of sucrose to isomaltulose, a new generation of sugar. From the previous study, palI gene from Klebsiella oxytoca was succesfully isolated from sapodilla fruit (Manilkara zapota). The full-length palI gene sequence of Klebsiella oxytoca was cloned in E. coli DH5α. The deduced amino acid sequence shows 498 residues which includes conserved motif for sucrose isomerisation 325RLDRD329 and 97% identical to palI gene from Klebsiella sp. LX3 (GenBank:AAK82938.1). This fragment was succesfullly ligated into the expression vector pET-32b using overlap-extension PCR and cloned in Escherichia coli BL21 (DE3) pLysS. DNA sequencing result shows that palI gene of Klebsiella oxytoca was inserted in-frame in pET-32b. This is the first report on cloning of palI gene from Klebsiella oxytoca.