Research Journal of Biotechnology

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Research Journal of Biotechnology





Genetic characterization of Awassi (Naeimi) sheep in Saudi Arabia based on microsatellite markers

Ahmed H. Mahmoud, Faisal Abou-Tarboush, Ahmed Rady, Nabil Amor, Khalid Alanazi and Osama B. Mohammed

In Saudi Arabia, the sheep production systems are mainly managed under farmer production systems, so their genetic management has led to increased homozygosity and hence productivity loss. The aim of this study was to genetically characterize 47 individuals of Awassi (Naeimi) sheep breed using a panel of eighteen microsatellite markers. Blood samples were used to extract genomic DNA followed by polymerase chain reaction (PCR). Results showed that 159 alleles were generated by the 18 microsatellites loci with an average of 8.833 alleles per locus. The DYMS1 SSR marker showed the highest number of alleles per locus (14) while OarAE129 showed the lowest (3). The expected heterozygosity (He) ranged from 0.541 to 0.860 with an average of 0.755 whereas the average observed heterozygosity (Ho) was 0.800 with a range of 0.319 to 1.000. The mean effective allele number was 4.643 which ranged from 2.177 to 7.219. Seven out of the 18 loci encountered significant departure from Hardy Weinberg Equilibrium (p < 0.05). The mean of Shannon index was 1.691 which ranged from 0.896 to 2.223. The results indicated the utility of using microsatellite loci in studying the genetic diversity in sheep populations of Saudi Arabia.

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Vermi products and biodegradable superabsorbent polymer improve physiological activities and leaf nutrient contents of gerbera

- Verma A.K., Sindhu S.S., Anand P., Singh A., Chauhan V.B.S. and Verma S.K.

Consistent with the fact that limited water and nutrient availabilities adversely impact the growth of potted plants, we investigated the effects of vermi-products and Pusa Hydrogel on physio-biochemical relations of gerbera cv. Yosemite. The potting media having CPV (Cocopeat: Perlite: Vermiculite in 4:1:1 ratio) and the soils amended with vermicompost (VC), pusa hydrogel (PHG) and/or horn bio-manure (HBM) had significant positive effects on plant physiological parameters that greatly influence growth and flower yield. Amelioration of growing media with 20% VC, 0.25% PHG and 1.0% HBM and the application of 20% vermiwash (VW) proved superior over other treatments in terms of marked improvements in leaf chlorophyll content, gas exchange characteristics and mineral acquisition by gerbera plants. In most of the cases, incremental additions of amendments and supplemental VW spray positively influenced these traits. Notably, 20% VW almost invariably proved superior over its lower (10%) concentration with regard to the parameters studied. Correlation study revealed that photosynthesis significantly increased with increase in stomatal conductance. Similarly, plants having high relative water content in leaves also maintained higher chlorophyll levels. The beneficial effects of soil substitution and/or enrichment with different media seemed partly due to favourable changes in the physico-chemical properties of the growing media. It also appeared that conjunctive use of amendments and supplementary VW applications acted synergistically to enhance the plant performance. These findings lead to the conclusion that incorporation of vermi-products and superabsorbent polymers in conventionally used media may prove beneficial for gerbera plant growth and flower yield.

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Discrimination of plant beneficial rhizosphere microbes by Amplified Ribosomal DNA Restriction (ARDRA) for molecular taxonomy

Jibu Thomas and Reena Josephine C.M.

A variety of microorganisms have been reported to promote the growth of the plants positively. Such beneficial bacteria that affect the growth of plants hastily are collectively called as plant growth promoting rhizobacteria. Identifying them accurately either taxonomically and characterising them for their potential are always cumbersome. Less discrimination capacity and high consumption of time in analysis are the setbacks for identification by phenotypic methods. Analysing the plant beneficial property of these strain would also demand taxonomical positioning of the strain for any further commercial applications. In the present study, beneficial plant growth promoting microorganisms (PGP) isolated from the rhizhospere of agricultural fields were attempted for characterization by various methods. Various biochemical tests were performed to confirm their PGP traits. These unknown culturable organism were amplified using Amplified Ribosomal DNA Restriction (ARDRA) to generate a fingerprint specific to these organisms before they were identified using molecular markers. A dendrogram based on ARDRA profile pattern was constructed using unweighted pair group method (UPGMA) to position them accordingly. The phylogenetic distances existed among the strains were thus established by this method. Organisms exhibiting clear discrimination in the ARDRA fingerprint were subjected to 16rRNA sequencing and further analysed to position them taxonomically by better means. Sequences generated for the novel strains were submitted to NCBI.

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In vitro production of disease free planting material of ginger (Zingiber officinale Rosc.) - A single step procedure

Thakur Manisha, Sharma Vishal and Kumari Garima

A protocol for in vitro multiplication of ginger was developed for the production of disease free planting material in a single step procedure. Sterilized vegetative buds excised from stored rhizomes were cultured on MS medium supplemented with 1.0 mg/l BA and 0.1 mg/l NAA for shoot differentiation. This medium was found to be the best for direct shoot regeneration from the vegetative buds taken as explants. Well-developed shoots were multiplied on MS medium supplemented with 1.0 mg/l BA, 0.1 mg/l NAA and 0.1 mg/l GA3 and showed enhanced multiplication rate of 1:14 after 4th sub-culturing. Rooting was observed in the shoot multiplication medium itself after 2-3 passages of sub-culturing, thus eliminating the additional step of rooting during micropropagation. In vitro grown plants were successfully hardened with 90% survival by transferring to sterilized potting mixture comprising of sand, soil and FYM (1:1:1).

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Bioremediation for uprooting heavy metals from contaminated groundwater: A case study (SIPCOT, PERUNDURAI, ERODE)

Durgadevagi S., Annadurai R. and Senthilkumar P.

The study area SIPCOT industrial estate is surrounded by 71 textile industries. Due to improper disposal of effluents, the groundwater in the area is highly contaminated and the rate of heavy metals such as Cu, Fe, Mn, Ni and Zn present in the groundwater is extremely high that leads to serious health issues and hence, removal of such heavy metals from groundwater is most important. Many conventional technologies have been used for removal of such heavy metals, but majority of them will cost high and not so effective to obtain potential output. Currently, a limitless exhibit of living materials, especially bacteria, algae and yeasts, work worthy for removal of heavy metals. In addition, bioremediation is cost-effective and output potential is comparatively high. In this case study, bacteria strains were isolated from groundwater samples and fifteen isolates were further studied based on its tolerance to various heavy metals. Based on the morphological and biochemical analysis the bacterial isolates were identified as Bacillus sp., Pseudomonas sp. and Staphylococcus sp. ICP-OES based analysis revealed that the bacterial isolates exhibit a range of resistance to Zn, Cu, Ni.

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Molecular analysis of beta satellites associated with okra yellow vein mosaic disease in Punjab (India)

Kaur Gurpreet, Sarao Navraj Kaur, Sharma Abhishek and Khan Irfan

Beta satellites from infected okra samples, exhibiting symptoms of yellow vein mosaic disease were analyzed for nucleotide similarity and phylogenetic relationship. The sequences showed high level of nucleotide similarity with the beta satellites associated with crops other than okra. Isolate from Moga showed 92.5% nucleotide similarity with Chilli leaf curl virus associated beta satellite whereas isolate from Gurdaspur was found 93.8% similar to Luffa puckering and leaf distortion beta satellite and okra isolate from Ludhiana with Ludwigia leaf curl disease associated beta satellite (90%). The results indicated the association of different beta satellites with yellow vein disease of okra in this region and spotlighted that these beta satellites are expanding their adaptation range by making new disease complexes with the viruses already present in the region.

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Cichorium intybus L. accords hepatoprotection in Streptozotocin induced diabetes mellitus in Wistar rats

Ramya Devi K. T.

Hepatoprotective potential of a Cichorium intybus (CIE) aqueous extract in streptozotocin (STZ)-induced diabetes mellitus in vivo is evaluated. The hepatoprotective assessment was done by estimating biochemical parameters such as alanine transferase (ALT), aspartamine transferase (AST) and alkaline phosphatase (ALP) and a histopathological study of liver was also performed. Administration of streptozotocin (60mg/kg) with and without CIE in experimental animals was studied for biochemical and histological deterioration. The groups administered with glibenclamide (5mg/kg body weight) and aqueous extract of CIE (250mg/kg body weight) showed a significant reduction in serum ALP, ALT and AST with 126.8 ± 5.2, 29.47 ± 2.0 and 42.3 ± 3.1 IU/mL levels respectively, compared to the 60mg/kg body weight groups treated with STZ with results of 224.4 ± 7.2, 48.1 ± 3.8 and 72.8 ± 3.5 IU/mL respectively. Histopathological observations of rat liver treated with CIE showed significant protection against hepatic damage compared with the diabetic control with respect to hepatic lobules, uniformity in cytoplasm distribution and infiltration. Correlation of the biochemical and histopathological parameters of the groups administrated with CIE (250mg/kg of body weight) showed maximum hepatoprotective potential.

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Molecular Analysis of the Bacterial Communities from Tannery Contaminated Sites in Riyadh, Saudi Arabia

Majed Albokari, Amr Arishi and Ashraf Essa

A major problem of the tanning industry is the disposal of chromium-contaminated wastes which have a deleterious effect on the environment. Inductively Coupled Plasma Mass Spectrometer analysis of tannery dumping soils (Riyadh, Saudi Arabia) showed the presence of high concentrations of chromium (268–297 ppm) and strontium (255–435 ppm). 454 pyrosequencing was applied to characterize the bacterial communities in four contaminated sites and a reference site. A total of 5,862 bacterial clones were assembled through 16S rRNA gene clone libraries. Phylogenetic analysis at the species level demonstrated contrasting distributions, interrelationships and shared species in all sites including the reference site. The chromium-contaminated sites were dominated by bacterial phyla Proteobacteria, Bacteroidetes and Firmicutes while other genera such as Chloroflexi, Acidobacteria, Gemmatimonadetes, Cyanobacteria and Nitrospirae were not detected. The bacterial genera Halomonas, Proteiniphilum, Alkaliphilus and Marinilabiaceae were identified only in the contaminated sites revealing their capability to tolerate and resist chromium toxicity. A vast variation in bacterial species was observed where Halomonas xinjiangensis was found at a considerable percentage in the chromium polluted soils. These findings revealed that long-term chromium stress in the tannery or dumping soil resulted in community shifts towards a dominance of chromium-resistant bacterial populations.

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Germplasm conservation of Clitoria ternatea L. through storage of encapsulated shoot tips and maintaining nodal segments under slow growth conditions

Dixit Jyoti, Singh Jayanti and Tiwari Kavindra Nath

The presence of 3% sodium alginate+100µM calcium chloride was found most suitable for Clitoria ternatea synthetic seed production and subsequent conversion of encapsulated shoot tips into plantlet. MS medium used for encapsulation of shoot tips showed highest conversion frequency (90±7.0%) on MS medium supplemented with 1.0mg/l BAP. Encapsulated shoot tips stored for 75 days at 4ºC showed maximum conversion frequency (52±3.2%) on MS + 1.0mg/l BAP + 0.1mg/l IBA. Nodal segmented stored on ½ MS medium supplemented with 4% sorbitol for 12 months showed highest conversion frequency (65%) and maximum number of shoots per explants on MS medium containing 0.5mg/l BAP. Elongated shoots showed best rooting on MS + 0.2 mg/l IBA. Plants derived from short term stored synthetic seeds and nodal segment showed normal flowering and growth.

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Molecular Characterization of Microbial Diversity in Ecologically Unique Niche Vedanthangal Lake by Ribotyping

Bindusree G., Jayasakthi E. and Iyappan S.

Vedanthangal Lake Bird Sanctuary is located in Kancheepuram District, Tamil Nadu, India. The sanctuary is about 75 km from Chennai, located south of Chengalpattu. Since this area is an ecologically unique niche, a study on microbial diversity of Vedanthangal Lake was carried out. Water samples from the lake were collected and pure cultures were isolated by spread plating. Fifty organisms were isolated. Based on morphology, 20 were selected for identification. Organism identification was performed by amplification and sequencing of 16S rDNA using fD1 and rP1 primers. When compared with the prototype strain in the GenBank database from NCBI, one sample among the identified samples showed 100% similarity; six samples showed 99% similarity, two samples showed 98% similarity and one sample showed <95% similarity. A phylogenetic tree was constructed that showed the phylogenetic distance relationship among the samples. Most of the species belong to Bacillus sp. and this unique site may be a potential site for screening for industrially important microbes.

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Gas Chromatography analysis of different plant parts of Phlogacanthus thyrsiflorus Nees

Singh Shweta, Banerjee Madhuparna and Kumar Manish

The species Phlogacanthus thyrsiflorus Nees, a plant native to northeast India, has been used by the traditional healers for different therapeutic purposes. The objective of this work was to investigate the secondary metabolites present in different parts of P. thyrsiflorus using gas chromatography-mass spectrometry(GC-MS). The GC-MS analysis of the methanolic extract of leaves, flowers and roots of this plant using NIST-14 MS library revealed the presence of several secondary metabolites of medicinal importance. In root extract, Lupeol was found in maximum (34.43%) followed by trans cinnamic acid (9.38%). In leaf extract, neomenthol was found highest (26.63%) followed by n-hexadecanoic acid (22.95%) whereas flower extract was found rich in 2, 3 - Dihydro - 3, 5 -dihydroxy - 6 - methyl - 4H - pyran - 4 - one (DDMP) (31.23%). A few of these chemicals possess medicinal properties comparable to the therapeutic uses of the plant parts by the traditional healers.

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Molecular Marker Techniques used in Plant Genomics

Mutthanthirige Don Lalith Chandana Nishantha and Song Weining

DNA fingerprinting of species has been structured for the recognition of taxa to advance relevant plant breeding strategies through management of germplasm. Several kinds of DNA markers have proved their utility for genetic variation analysis in various species both at inter and intra specific level. The development and use of molecular markers for the detection and exploitation of DNA polymorphism have been recognized as an important area of study under molecular genetics. Detection and analysis of genetic variation can help us to understand the molecular basis of various biological phenomena in plants. Genetic or DNA based marker techniques are regularly being used in ecological, evolutionary, taxonomical, phylogenic and genetic studies of plant sciences. The technical advancements and genome based discoveries have led to the enhancement of molecular marker techniques. The presence of different molecular markers and characteristically different principles, methodologies and functions require cautious attention in selecting one or more of such techniques. According to the kind of study to be carried out, researcher can choose among the variety of molecular techniques, each of which combines at least some desirable properties. This review describes major molecular markers and related techniques used in plant genomics.

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