Genomic DNA extraction from sapwood of Pinus roxburghii for polymerase chain reaction studies

A method for extraction of genomic DNA from sapwood tissues of mature tall trees of Pinus roxburghii, where collection of needle tissues is extremely difficult has been standardized. The extracted DNA was comparable to that obtained from the needle tissue in terms of yield and purity. The yield of extracted DNA ranged from 6.98 to 19.668 μg / 100 mg tissue and A260 / A280 ratio ranged from 1.70 to 1.87. The polymerase chain reaction (PCR) amplification of the DNA extracted from sapwood tissue using random amplification of polymorphic DNA (RAPD), inter simple sequence repeat (ISSR) and simple sequence repeat (SSR) markers was similar to that of DNA extracted from the needle tissues.Keywords: Pinus roxburghii, DNA extraction, sapwood, random amplification of polymorphic DNA (RAPD), inter simple sequence repeat (ISSR), simple sequence repeat (SSR)African Journal of Biotechnology Vol. 12(15), pp. 1732-173

Identification of allele specific primers in chir pine (<em>Pinus roxburghii</em> Sarg.) through data mining

538-540Pinus roxburghii Sarg. commonly known as Chir pine, is a commercially exploited species for resin in India. Resin is a phenotypic trait which is expressed only in mature trees. Due to this a large number of trees are cut every year which pose a serious ecological threat. Recent advancements in bioinformatics and molecular biology have enabled scientists to identify high resin yielding pine genotypes at nursery stage using molecular markers. In the present study, investigation, 337 terpene coding sequences from the database were analyzed using data mining and 45 microsatellites allele specific primers were designed for marker assisted selection. Out of these 45 primers, only 22.2% primer pairs showed polymorphism within the 53 genotypes of Chir pine examined. A wide range of fragment size was observed from 100 - 600 bp. PCR amplification using allele specific markers produced a total of 22 bands, out of which 14 were found to be polymorphic. The total number of bands amplified per marker varied from of 1 to 3 with an average of 1.4. Genetic divergence in terms of percent polymorphism ranged from 50 to 100% with a mean of 63.3% per marker

Response of Populus deltoides Bartr. ex Marsh genes under elevated CO2 through Next-Generation Sequencing (NGS)

The impact of climate change has attracted considerable attention globally. Atmospheric Carbon Dioxide (CO2) is expected to increase to 900 μmol mol-1 from present level of 400 μmol mol-1 by the end of 21st century. CO2 is a greenhouse gas that leads climate change have significantly affected structure and function of the terrestrial ecosystem, global carbon, water balance, and also crop productivity. These responses of the plant appear by altering gene expression pattern of different genes involved in anabolic and catabolic processes. We have conducted a study to see the response of genes to elevated CO2 inside open top chambers on Populus deltoides. Onemonth- old ramets were exposed for 180 days to treatment (CO2 800 μmol mol-1) and control (CO2 ~400 μmol mol-1). After completion of treatment, leaf tissues were outsourced to Sci-genome for transcriptome sequencing. This study demonstrated, higher (1754) number of transcript expression in treatment (119,306) compared to control (121,060). Differential gene expression analysis shown 1951 transcripts were down regulated while 2603 transcripts up regulated and 159,982 transcripts have no significance in treatment. Our results show that plants growing in an environment where atmospheric CO2 is higher may alter plant adaptation, productivity, vegetation and ecosystem health by changing; the first, number of genes and second, altering gene expression patterns. Such behavior may be a good indicator of developing adaptation strategies of the plant

Genomic DNA isolation and identification of chloroplast microsatellite markers in <i>Asparagus racemosus </i>Willd.<i> </i>through cross-amplification

33-38Cross-species amplification of microsatellite loci is a time saving as well as a cost-effective approach for developing locus specific markers for new species. In an attempt to reveal the genetic variation in different accessions of Asparagus racemosus, chloroplast microsatellite primer pairs developed for Acorus calamus (Acoraceace) were examined for cross-species amplification and validation in A. racemosus. Out of the 18 microsatellite primer pairs screened, 5 i.e. 27.77% (AC-03; AC-05; AC-09; AC-13 and AC-17) yielded good cross-species amplification across 20 different individuals of A. racemosus. These cpSSR markers comprise 1-dinucleotide repeat type; 2-trinucleotide and 2-tetranucleotide repeat types. The product size of the amplified cpSSR primers ranged between 180 and 337 bp. All the 5 cross-amplified cpSSR markers were found polymorphic across the 20 individuals of A. racemosus. Besides this an easy and competent protocol for the extraction of high quality genomic DNA in A. racemosus for the PCR-based microsatellite marker analysis has been also reported. The DNA extraction protocol involved a modification of CTAB procedure given by Stange et al, which includes the use of high concentrations of polyvinyl pyrrolidone (PVP), addition of 8 M lithium chloride in extraction buffer; a repeated chloroform:isoamyl alcohol step and washing of DNA pellets with the wash buffer and with the 80% ethanol. The developed protocol yielded approximately ~77.30 μg DNA per 100 mg plant tissue with the purity ratio of ~1.85 at A260/A280 nm wavelength. Following the protocol and using the primers, genetic diversity analysis in A. racemosus was carried out

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 August 2010-30 September 2010

This article documents the addition of 229 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Acacia auriculiformis × Acacia mangium hybrid, Alabama argillacea, Anoplopoma fimbria, Aplochiton zebra, Brevicoryne brassicae, Bruguiera gymnorhiza, Bucorvus leadbeateri, Delphacodes detecta, Tumidagena minuta, Dictyostelium giganteum, Echinogammarus berilloni, Epimedium sagittatum, Fraxinus excelsior, Labeo chrysophekadion, Oncorhynchus clarki lewisi, Paratrechina longicornis, Phaeocystis antarctica, Pinus roxburghii and Potamilus capax. These loci were cross-tested on the following species: Acacia peregrinalis, Acacia crassicarpa, Bruguiera cylindrica, Delphacodes detecta, Tumidagena minuta, Dictyostelium macrocephalum, Dictyostelium discoideum, Dictyostelium purpureum, Dictyostelium mucoroides, Dictyostelium rosarium, Polysphondylium pallidum, Epimedium brevicornum, Epimedium koreanum, Epimedium pubescens, Epimedium wushanese and Fraxinus angustifolia