Isolation of high quality genomic dna from dryobnanalops beccarii dyer tissues rich in camphor

The isolation of high-quality intact genomic DNA is obviously difficult in a variety of plant species due to the presence of high levels of polysaccharides, polyphenolics and other secondary metabolites. These contaminants will co-precipitate with DNA during DNA isolation and purification processes, and therefore, resulting in a brownish DNA pellet that is unsuitable for downstream applications. Most plant genomic DNA isolation protocols for various plant tissues have been established; however these protocols are inefficient in yielding high-quality amplifiable genomic DNA especially from camphor containing timber species, Drynobanalops beccarii Dyer. In this project, the total genomic DNA of Drynobanalops beccarii was successfully isolated using a re-optimized CTAB method based on the Murray & Thompson (1980) and Doyle & Doyle (1990). The modifications include: 1) using 1% =-mercaptoethanol and 2% PVP (Mr 40000) in the extraction buffer, 2) sample incubation time, 40 minutes at 65°C; and 3) DNA precipitation at room temperature (25°C). The isolated DNA pellet was in transparent colour and yielded amplifiable genomic DNA for RAPD-PCR

An Optimized DNA Extraction Protocol For Isolation Of High Quality Genomic DNA From Camphor Containing Timber Tree Species, Dryobalanops Beccarii Dyer

Isolation of high-quality genomic DNA from Dryobalanops beccarii is obviously difficult due to the existence of large amounts of camphor and other secondary metabolites. These contaminants will co-precipitate with DNA during DNA isolation and purification processes, and therefore, resulting in a brownish DNA pellet that is unsuitable for downstream applications. Many DNA isolation protocols are available for various plant tissues; however these protocols are inefficient in yielding high-quality amplifiable genomic DNA especially from camphor containing timber tree species. A CTAB based protocol has been optimized for isolating genomic DNA from camphor containing timber tree species. Key steps include: 1) using 1% β-mercaptoethanol and 2% PVP 40 (Mr 40,000) in the extraction buffer; 2) sample incubation time, 40 minutes at 65°C, and 3) DNA precipitation at room temperature (25°C). The isolated DNA pellet was transparent colour and the purified genomic DNA is suitable for PCR amplification. Wei-Seng Ho, Kit-Siong Liew, Shek-Ling Pan

Development, Polymorphism and Cross-Species Transferability of Genomic SSR Markers in Duabanga Moluccana, an Indigenous Tree Species from Sarawak

In this study, we used ISSR-suppression methods to develop a set of SSR markers for Duabanga moluccana. It is an indigenous fast growing tropical tree species. A total of 44 SSR regions were identified and specific primer pairs were designed. The SSR motifs contained perfect compound with 24 (54.5%) occurrences, followed by the imperfect compounds with 8 (18.2%), simple perfect with 8 (18.2%) and the simple imperfect repeats with 4 (9.1%). The newly identified SSR markers were characterized by screening 20 individuals of D. moluccana seedlings. Among 43 primer pairs tested, 25 (58.1%) SSR markers amplified the desired PCR products and 115 alleles were detected. The number of alleles per locus ranged from 2 to 8, with a mean value of 4.60. Polymorphism Information Content (PIC) values ranged from 0.225 to 0.792, with an average of 0.604. A success rate of transferability of D. moluccana SSR markers varied, ranging from 84% in Duabanga grandiflora, 36% in Neolamarckia cadamba, 24% in Canarium odontophyllum and 28% in Shorea parvifolia. These SSR markers herein could be used to generate useful baseline genetic information for effective selection of plus trees, provenance trials and establishment of forest Seed Production Areas (SPAs) of D. moluccana in the selected forest reserves for tree plantation and improvement activities. Besides, the transferability of the newly developed SSR markers across a range of species and genera suggests their potential usefulness for a variety of population genetic studies

Determination of genetic diversity and genetic relatedness using ISSR markers in Duabanga moluccana

Duabanga moluccana or locally known as Sawih is a widely known forest tree species for its multi-purpose timber and other natural products such as fibers. Genetic diversity is important for the maintenance of the viability and the adaptive potential of populations and species. This information will be a basis for establishing tree improvement programme and for management or conservation of natural communities. The present study was aimed to assess the genetic diversity and genetic relatedness of D. moluccana trees collected from three natural forests in Sarawak, namely, Mukah, Tatau and Niah using inter simple sequence repeat (ISSR) markers. A total of 73 loci from 90 individuals were successfully amplified using three selected ISSR primers, namely, (AC)10, (ACC)6G and ACG(GT)7. The Shannon’s diversity index showed that D. moluccana trees in Mukah (0.499) was the most diverse population compared to Tatau (0.380) and Niah (0.330). Neighbor joining tree was also constructed to determine the genetic relationship among the three D. moluccana populations. The populations were completely clustered into three main groups, in accordance to their corresponding population and origin. Based on the results, it implies that D. moluccana trees are genetically diverse and related both within and among populations

From Conservation To Innovation Building Research Capacity For Planted Forest Development In Sarawak

The increase in global demand for wood requires increase in forest productivity. The alternative is to farm trees in plantations composed of fast-growing species with short rotation cycle (6-8 years). The rationale is that natural forests at the most produce about 3m3/ha/yr of commercial timber, whereas plantations can produce annually from 10m3/ha of hardwoods to 30m3/ha of softwoods and thus, decrease the effects of human pressure on our ecosystems while increasing the competitiveness of Sarawak‘s forest industry. This is in line with State Government‘s aspiration to establish one million hectares of planted forests by year 2020 to meet the increasing demand from both domestic and international markets for raw materials. It is estimated at least 30 million seedlings are required for annual planting or reforestation programmes. In this regard, the forest genomics research will help respond to the need to develop adequate tools that enable us to produce quality planting materials that are of faster growth, high-yield and high wood quality, and also adapted to local conditions, so that we may achieve economic benefits of great significance. Realizing the needs, we have centered our research on the development of tools via biotechnological innovations for tree breeders. We have successfully developed: 1) an array of highly informative and polymorphic DNA markers specific for identifying the genetic makeup of two fast growing indigenous tree species, i.e. Kelampayan and Sawih; 2) the one step ‗Touchincubate- PCR‘ approach for preparing plant tissues for high throughput genotyping, and 3) a genomic resource database, aka CADAMOMICS (10,368 ESTs) for wood formation in Kelampayan via high-throughput DNA sequencing. These tools will greatly facilitate the selection of quality planting materials for planted forest development in Sarawak as well as longterm tree improvement activities by integrating genomics into our breeding programme via association mapping. The overall benefit of genomics application to tree improvement programme will be in terms of greater certainty in the outcome of results, specifically the performance of the forest plantations, as well as the savings in time and cost in the production and supply of quality planting materials

Development and polymorphism of simple sequence repeat (ssr) dna markers for Duabanga moluccane blume

Duabanga moluccana Blume (or locally known as Sawih) is an indigenous fast growing tree species that has been selected for planted forest development in Sarawak. It possesses great commercial values in production of various wood products. Therefore, the development of an array of simple sequences repeats (SSRs) or micro satellite DNA based-markers is absolutely necessary to study the genetic makeup of this species. These molecular markers had been rapidly used in the selection of plus trees, quality control in seed production, investigation of population genetics and conservation management. Isolation of high-quality genomic DNA from D. moluccana is vital for molecular marker development as well as other downstream application) In the present study, an improved method of total DNA isolation from D. moiuccana was established. These modifications include: 1) precipitating DNA with '/3 volwne of 5 M NaCI together with isopropanol; 2) using I% ~-mercaptoethanol in the extraction buffer; 3) sample incubation time of 40 minutes at 65 DC, and 4) adding a CIA extraction step. By using this method, the isolated DNA was suitable for amplification and restriction digestion analysis. In this study, we used ISSR-suppression method to develop simple sequence repeats (SSRs) markers for D. moiuccana. Overall, 44 SSR regions were identified. The microsatellite motifs contained simple perfect and simple imperfect microsatellites constituted by di-and trinucleotides and, perfect/imperfect compounds. The most nwnerous class was the perfect compound with 24 (54.5%) occurrences, followed by the imperfect compounds with 8 (18.2%), simple perfect with 8 (18.2%), and the simple imperfect repeats with 4 (9.1 %). Majority of the dinucleotide microsatellites found in the Sawih genome was AG/GA/CT/TC repeats (83.3%) followed by ACICAITG/GT repeats (16.7%). Primer pairs were designed for 43 SSR regions. The newly identified SSR markers were characterized by screening DNA templates from 20 individuals of D. moluccana seedlings. Among 43 primer pairs, 25 (58.1 %) SSR markers amplified the expected fragments size while 17 (39.5%) produced unexpected PCR products or multiple bands. One primer did not generate amplification products in most of the Sawih samples. A total of 115 alleles were detected across 25 loci analysed. The number of alleles per locus ranged from 2 to 8, with a mean value of 4.60. Polymorphism Information Content (PIC) values ranged from 0.225 to 0.792, with an average of 0.604. A success rate of transferability of D. moluccana microsatellite markers varied, ranging from 84% in Duabanga grand~flora, 36% in Neolamarckia cadamba, 24% in Canarium odontophyllum and 28% in Shorea parvifolia. The development of an array of microsatellite markers herein could be applied to generate useful baseline genetic information for effective selection of plus trees, provenance trials, and establislunent of forest seed production areas (SPAs) of D. moluccana in the selected forest reserves for tree plantation and improvement activities. Besides, the transferability of the newly developed microsatellites markers across a range of species and genera suggests their potential usefulness for a variety of population genetic studies

Struggle for democracy : Sung Chiao-jen and the 1911 Chinese Revolution

The 1911 revolution was a momentous event in Chinese history. It overthrew the 2,000-year-old monarchical system, and tried to establish a democratic republic in its place. The failure of this first attempt to Westernise China, combined with the other frustrations of the young intellectuals who engineered the revolution, contributed in large measure to China{u2019}s disenchantment with democracy, and to her subsequent intense commitment to Communism. This book, set against the background of events, is a study of the handful of young revolutionaries who nurtured and united radical feeling in China so as to bring about the revolution; in particular, it is a study of Sung Chiao-jen, revolutionary leader, idealist, and intellectual, who was assassinated at the age of thirty at Shanghai in 1913. The beliefs and aspirations, the struggle for democracy, the disillusionments of Sung Chiao-jen and his fellow revolutionary leaders, provide the necessary background of understanding for judgments about China{u2019}s last sixty years. It is a story that everyone interested in China will want to read

Sung Chiao-Jen : a study of his revolutionary and political career

This is a study of Sung Chiao-jen also known as Tun-ch'u or, pen name Yu-fu (1882-1913). The aim is twofold. Firstly, it is to tell the story of a man who played an intimate role in the Chinese Revolution of 1911 and sacrificed his life for the cause of democracy. Secondly, it is hoped that such a study will help to elucidate the nature of this revolution

Development and characterization of microsatellite markers in sawih tree (Duabanga moluccana Blume) using ISSR-suppression PCR techniques

Duabanga moluccana (or locally known as sawih) is an indigenous fast growing tropical tree species that confers various advantages for the timber industry and for planted forests development. In this paper, we isolated and characterized 8 polymorphic microsatellite markers from the D. moluccana genome using ISSR-suppression PCR techniques. The number of alleles and PIC values ranged from 3 to 8 alleles per locus and from 0.488 to 0.792, respectively. Three microsatellite loci were deviated from Hardy-Weinberg equilibrium (P<0.05). The transferability rate ranged from 24 to 100 % among the three indigenous tree species tested. This indicates that the newly developed microsatellite markers would be useful tools for population genetic studies on D. moluccana and other indigenous tree species

Development and characterization of microsatellite markers in sawih tree (Duabanga moluccana Blume) using ISSR-suppression PCR techniques

Duabanga moluccana (or locally known as sawih) is an indigenous fast growing tropical tree species that confers various advantages for the timber industry and for planted forests development. In this paper, we isolated and characterized 8 polymorphic microsatellite markers from the D. moluccana genome using ISSR-suppression PCR techniques. The number of alleles and PIC values ranged from 3 to 8 alleles per locus and from 0.488 to 0.792, respectively. Three microsatellite loci were deviated from Hardy-Weinberg equilibrium (P<0.05). The transferability rate ranged from 24 to 100 % among the three indigenous tree species tested. This indicates that the newly developed microsatellite markers would be useful tools for population genetic studies on D. moluccana and other indigenous tree species