Easy DNA Extraction Protocol for Ricinus communis Seeds | InformativeBD

Ajmal Iqbal,  Waqar Ahmad,  Asaf Khan,  Murad Khan, and Mohammad Nisar, from the institute of Pakistan,. wrote a Research article about, Easy DNA Extraction Protocol for Ricinus communis Seeds. Entitled, User friendly DNA isolation protocol optimized for Ricinus communis L. seeds. This research paper published by the International Journal of Biosciences | IJB.  an open access scholarly research journal Biosciences. under the affiliation of the International Network For Natural Sciences| INNSpub. an open access multidisciplinary research journal publisher.

Abstract

A protocol was developed to isolate high quality genomic DNA from the seeds of Ricinus communis L. (caster) without using liquid nitrogen.The DNA extraction buffer used in this novel protocol constitutes SDS (1%), Tris (1.21%), NaCl (0.58%), EDTA (0.32%), 0.12% and β–Mercaptoethanol witha pH 8.5. In the protocol 0.09g of crushed seeds of caster bean, 600ul of DNA extraction buffer and and 500ul of phenol: chloroform: iso-amylalcohol with a ratio of 25:24:1 were used. The isolated DNA was amplified in Polymerase Chain Reaction using RAPD and SSR primer sets. The primer set successfully amplified the isolated DNA. Hence, the protocol is recommended as a user friendly novel protocol for DNA isolation from the castor beans seeds.

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Introduction

Ricinus communis L., commonly known as caster bean, is a member of the spurge family,Euphorbiaceae (Smith 1986). Ricinus is indigenous to tropical Asia and Africa, but today it is cultivated for seed oil throughout the tropical and subtropical regions of the world (Seo, 2011). The annual production of caster bean throughout the world is 460,000 tones. In Pakistan it is grown on 3204 ha and its annual production is 2089 tones, the average seed yield of caster bean in Pakistan is 652 kg ha-1 . This seed yield of Pakistan is very low (Anonymous, 2006). Oil derived from the seeds of caster has several potentials in many industries like medicine, and cosmetics (Akande, 2012). Castor bean seed oil is highly valued in several sectors of the chemical industry and is considered a bioenergy and phytoremediation resource in the subtropics (Baldanzi et al. 2005). Castor plant has a great drought tolerance due to deep root system with the ability to explore the deeper layers of soil, which helps increase aeration, water retention and distribution in soil (Embrapa et al., 2006).

The Preliminary Phytochemical study of Ricinus communis revealed the presence of steroids, saponins, alkaloids, flavonoids, and glycosides (Kang, et al., 1985). The leaves of the Ricinus communis contain flavonoids, tannins and phenol (Yadav RNS & Agarwala M. 2011; Ilavarasan et al., 2006).

The isolation of high-quality DNA is prerequisite for any molecular biology work because contaminants such as proteins, polyphenols and polysaccharides may interfere with enzymes, such as endonuclease (in blotting techniques) and Taq polymerase in Polymerase Chain Reaction (Ausubel et al., 1994). The phenols covalently bind to proteins and DNA, giving the DNA a brown colour and making it useless for most research applications (Katterman & Shattuck, 1983; Guillemaut & Drouard, 1992, Aljanabi et al., 1999). Polyphenol contamination of DNA makes it resistant to restriction enzymes (Katterman & Shattuck, 1983).

Several methods for extracting DNA for different plant are available (Doyle and Doyle et al., 1990;

Khanuja et al., 1999; Kumar et al., 2003 Islam et al., 2013). In Pakistan no work has been done on the extraction of DNA from Ricinus species. Therefore the aim of the present study is to develop simple DNA extraction protocol from the seeds of six different varieties of Ricinus communis without liquid nitrogen and for further genome characterization using RAPD and SSR primers.

Reference

Akande TO, Odunsi AA, Olabode OS, Ojediran TK. 2012. Physical and Nutrient Characterisation of Raw and Processed Castor (Ricinuscommunis L.) Seeds in Nigeria. World Journal of Agricultural Sciences 8, 89-95.

Aljanabi SM, Forget L, Dookun A. 1999. An improved and rapid protocol for the isolation of polysaccharide and polyphenol free sugarcane DNA. Plant Molecular Biology Reporter 17, 1–8. http://dx.doi.org/10.1023/A:1007692929505

Anonymous. 1948-1976. Council of Scientific and Industrial Research.The Wealth of India, New Delhi. 11.

Anonymous. 2006. Agri. Statistics of Pakistan. Ministry of Food, Agriculture and Livestock, Islamabad, Pakistan.

Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K. 1994. Current protocols in molecular biology. John Wiley and Sons, New York. 2.0.1–2.14.8.

Baldanzi  M,  Fambrini  M,  Pugliesi  C.  2005. Redesign  of  the  castor  bean  plant  body  plan  for optimal combine harvesting. Annals of Applied Biology 142, 299-306. http://dx.doi.org/10.1111/j.1744-7348.2003.tb00254.x

Barzegari A, Vahed SZ, Atashpaz S, Khani S, Omidi Y. 2010. Rapid and simple methodololgy for isolation of high quality genomic DNA from coniferous tissues (Taxus baccata). Molecular Biology Reporter 37, 833-837. http://dx.doi.org/10.1007/s1133-009-9634-z

Correa MP. 1984. Dicionario das Plantas Uteis do Brasil e das Exoticas Cultivadas. Ed. Imprensa Nacional Rio de Janeiro, 63.

Doyle JJ, Doyle JL. 1990. Isolation of plant DNA from fresh tissue. Focus 12, 13-15.

Embrapa-Empresa Brasileira de Pesquisa Agropecuaria 2006. Cultivo da mamona. Embrapa-CNPA, Algodao, Campina Grande, Sistemas de Produçao 4-2.

Feijao, RDO. 1963. Rícino. ElucidarioFitologico, ed. InstitutoBotanico de Lisboa, 3, 111-112.

Guillemaut P, Drouard ML. 1992. Isolation of plant DNA: fast, inexpensive, and reliable method. Plant Molecular Biology Reporter 10, 60–65. http://dx.doi.org/10.1016/j.gene.2010.08.009

Ilavarasan  R,  Mallika  M,  Venkataraman  S. 2006. Anti-inflammatory and free radical scavenging activity of Ricinus communis root extract Journal of Ethnopharmacology 103, 478 – 480. http://dx.doi.org/10.1016/j.jep.2005.07.029

Islam M, Ahmad H, Khan IA. 2013. An efficient protocol for DNA isolation from the genus Pyrus. International Journal of Biosciences 3, 122-127. http://dx.doi.org/10.12692/ijb/3.4.122-127

Kang SS, Cordell GA, Soejarto DD, Fong HHS. 1985. Alkaloids and flavonoids from Ricinuscommunis. Journal of Natural Products 48, 155–156. http://dx.doi.org/10.1021/np50037a041

Katterman FRH, Shattuck VL. 1983. An effective method of DNA isolation from the mature leaves of Gossypium  species  that  contain  large  amounts  of phenolic terpenoids and tannins. Preparative Biochemistry 13, 347–359. http://dx.doi.org/10.1080/00327488308068177

Khanuja SPS, Shasany AK, Darokar MP, Kumar S. 1999. Rapid isolation of DNA from dry and fresh samples of plants producing large amounts of secondary metabolites and essential oils.Plant molecular biology Reporter 17, 1 – 7. http://dx.doi.org/10.1023/A:1007528101452

Kumar A, Pushpangadan P, Mehrotra S. 2003. Extraction of high molecular weight DNA from dry root tissue of Berberis lycium suitable for RAPD. Plant Molecular Biology Reporter 21, 309a-309d http://dx.doi.org/10.1007/BF02772807

Li JF, Li L, Sheen J. 2010. Protocol: a rapid and economical procedure for purification of plasmid or plant DNA with diverse applications in plant biology. Plant Methods 6, 1. http://dx.doi.org/10.1186/1746-4811-6-1

Lodhi AM, Ye GN, Weeden NF, Reish. 1994. A simple and efficient method for DNA extraction from grapevine cultivars, Vitis species and Ampelopsis. Plant Molecular Biology Reporter 12, 6-13. http://dx.doi.org/10.1007/BF02668658

Puchooa D, Venkatasamy K. 2005.A protocol for isolation of DNA from Trocetia boutoniana. International Journal of Agriculture and Biology 7, 82-85.

Purohit AR, Verma PU, Patel NJ. 2012. Rapid and Efficient Procedure for Isolation of High Yielding DNA from Castor (Ricinus communis L.). International Journal of Scientific and Research Publications 2, 1–4.

Seo K, Lee G, Ma K, Hyun D, Park Y, Jung J, Lee S, Gwag J, Kim C, Lee M. 2011. Isolation and Characterization of 28 Polymorphic SSR Loci from Castor Bean (Ricinus communis L.). Journal of Crop Science and Biotechnology 14, 97 – 103. http://dx.doi.org/10.1007/s12892-010-0107-7

Smith AR. 1986. Euphorbiaceae. In E. Nasir and S. I. (edt.) Flora of West Pakistan. Department of Botany, Karachi University, Karachi. Pakistan 172.

Smyth RP. 2010. Reducing chimera formation during PCR amplification to ensure accurate genotyping. Genetics 469, 45-51. http://dx.doi.org/10.1016/j.gene.2010.08.009

Yadav RNS, Agarwala M. 2011. Phytochemical analysis of some medicinal plants. Journal of Phytology 3, 10-14.

Article source : User friendly DNA isolation protocol optimized for Ricinus communis L. seeds 

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Green Synthesis and Antimicrobial Potential of Silver Nanoparticles from Citrus aurantium | InformativeBD

R. Venkateshwari,  from the institute of India. R. Krishnaveni, from the institute of India. F. J. Jelin, from the institute of India. P. Bhuvaneswari, from the institute of India. T. Shanmuga Vadivu, from the institute of India. and G. Annadurai, from the institute of India.  wrote a Research Article about, Green Synthesis and Antimicrobial Potential of Silver Nanoparticles from Citrus aurantium. Entitled, Citrus aurantium bark, seeds, and leaves were used to synthesize and characterize silver nanoparticle and their antimicrobial activity was evaluated. This research paper published by the Journal of Biodiversity and Environmental Sciences (JBES).  an open access scholarly research journal on Biodiversity. under the affiliation of the International Network For Natural Sciences | INNSpub. an open access multidisciplinary research journal publisher.

Abstract

The green synthesis of silver nanoparticles has been proposed as an eco-friendly and cost-effective substitute for chemical and physical methods. The aim of this study was to synthesize and characterize silver nanoparticles using the peel extract of Citrus aurantium Bark, Leaf and Seed, and to determine the possible phytochemical constituents’ presence in the plant extracts that might be responsible for the synthesis. Citrus aurantium Bark, Leaf and Seed extraction was followed by phytochemical studies of secondary metabolites, FTIR analysis confirmation of functional groups analysis. Silver nanoparticles were synthesized through bio-reduction of silver ions to silver nanoparticles using Citrus aurantium Bark, Leaf and Seed and characterized using UV-Vis spectroscopy (Bark, Leaf and Seed), SEM (Bark and Leaf), XRD (Bark, Leaf and Seed) and FTIR (Bark Leaf and Seed). The FTIR analysis of the extract revealed the presence of functional groups like hydroxyl, carboxyl, carbonyl, amine, and phenyl with similar functional groups. The synthesized silver nanoparticle (AgNP) has displayed the characteristics of a UV-Vis spectroscopy band peak from 400–420 nm. The XRD analysis also confirmed that the nanoparticles synthesized are crystalline in nature. Based on the findings of this study, it is understood that the variety of natural compounds that are present in plant extracts of Citrus aurantium  Bark, Leaf and Seed can act as both reducing and stabilizing agents for the synthesis of silver nanoparticles. It is, therefore, concluded that Citrus aurantium Bark, Leaf and Seed extract can be potentially used for the large production of silver nanoparticles for several applications.

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Introduction

Due to their potential and potential applications in a variety of fields, including biomedicine, nanomedicine, agriculture, and biosensors, there has been an increase in interest in the synthesis of metallic nanoparticles such as zinc, silver, platinum, and gold in recent years (Tijjani Mustapha et al., 2023; Pirtarighat et al., 2019). Due to their high stability and low chemical reactivity compared to other metals, silver nanoparticles have been studied more than any other nanomaterial. Because of their unique and promising qualities, they are frequently employed as larvicidal, antibacterial, and anticancer agents (Tijjani Mustapha et al., 2023; Mustapha et al., 2022). Nonetheless, two distinct approaches are frequently used to synthesize them: the chemical and physical approaches. Typically, chemical or physical techniques such as micelle synthesis, sol process, chemical precipitation, hydrothermal method, pyrolysis, and chemical vapour deposition are used to create nanomaterials (Charusheela Ramteke et al., 2013; Leela and Vivekantandan, 2008). Certain techniques are simple and allow for the regulation of crystallite size through the restoration of the reaction environment. However, there are still issues with the product's overall stability and getting monodisperse nanosize using these techniques (Kowshik et al., 2002; Charusheela Ramteke et al., 2013). Furthermore, it has been discovered that a large number of conventional techniques are capitalintensive and inefficient in their use of materials and energy (Klaus-Joerger et al., 2001; Charusheela Ramteke et al., 2013).

A green method has recently been proposed to replace the methods that harm the environment, such as chemical and physical ones. The biological method also referred to as the green synthesis technique or method makes use of bacteria, fungi, and plants. Using plant extracts from different plant parts, including the peel, stem, leaf, root, and fruit, several studies have reported the green synthesis of silver nanoparticles (Charusheela Ramteke et al., 2013; Atharbi et al., 2018; Ayodele et al., 2020; Kokila et al., 2015). A flowering plant in the Rutacea family is called Citrus aurantium. The terms "bitter orange" and "key lime" are frequently used to describe them (Khan Pathan et al., 2012; Nur et al., 2016). It is one of the most widely used citrus species in Malaysia, where it is mostly utilized in traditional medicine and food. Its 3–5 m tall, spiky stem is its main feature. The citrus plant is spherical in shape, with leaves that are 3–5 cm thick and 5–9 cm long (Nur et al., 2016; Daigy, 2009; Mandal et al., 2009). Traditionally, tulsi leaves have been used to treat a variety of infections. It has been stated that the antibacterial activity stems from the components of essential oils, primarily the eugenols. The goal of this study is to create silver nanoparticles using Tulsi leaf aqueous extract. Additionally, in an effort to maximize antimicrobial action, we try combining the natural antibacterial properties of Tulsi extract with silver metal (Raghunan et al., 2011; Dubey et al., 2010; Baret et al., 2009).

Even though there have been a number of studies on silver nanoparticles, more thorough research is still needed on the environmentally friendly synthesis of silver nanoparticles utilizing plant extracts (GardeaTorresdey et al., 2003; Rafique et al., 2017). To our knowledge, no research has been done on the use of Citrus aurantium Bark, Leaf and Seed extract in the plant-mediated production of silver nanoparticle. Thus, identifying and characterizing the function of metabolites in the creation of silver nanoparticles constitutes the novelty of the current work. Considering the aforementioned, the purpose of this work was to use Citrus aurantium Bark, Leaf and Seed extract to synthesize and characterize silver nanoparticle and to identify potential phytochemical constituents present in the plant extracts that could be involved in the synthesis of the silver nanoparticle.

Reference

Albahadly Z, Albahrani R, Hamza A. 2019. Silver nanoparticles synthesized from Citrus aurantium L. & Citrus sinensis L. leaves and evaluation of antimicrobial activity. Journal of Global Pharma Technology 11(3), 71-75.

Amin M, Anwar F, Janjua MRSA, Iqbal MA, Rashid U. 2012. Green synthesis of silver nanoparticles through reduction with Solanum xanthocarpum L. berry extract: Characterization, antimicrobial and urease inhibitory activities against Helicobacter pylori. Int. J. Mol. Sci 13, 9923-9941.

Ayodele M, Chikodiri V, Adebayo-Tayo BC. 2020. Green synthesis and cream formulations of silver nanoparticles of Nauclea latifolia (African peach) fruit extracts and evaluation of antimicrobial and antioxidant activities. Sustain. Chem. Pharm 15, 100197.

Bar H, Bhui DH, Sahoo PG, Sarkar P, De PS, Misra A. 2009a. Green synthesis of silver nanoparticles using latex of Jatrapha curcas. Colloids Surf A Physicochem Eng Asp 339, 134–139.

Bar H, Bhui DK, Sahoo GP, Sarkar P, Pyne S, Misra A. 2009b. Green synthesis of silver nanoparticles using seed extract of Jatropha curcas. Colloids Surf A Physicochem Eng Asp 348, 212–216.

Charusheela R, Tapan C, Bijaya KS, Ram-Avatar P. 2013. Synthesis of silver nanoparticles from the aqueous extract of leaves of Ocimum sanctum for enhanced antibacterial activity. Journal of Chemistry 278925, 1-7.

Daizy P. 2009. Biosynthesis of Au, Ag and Au–Ag nanoparticles using edible mushroom extract. Spectrochimica Acta Part A 73, 374–381.

Deshpande R, Bedre MD, Basavaraja S, Sawle B, Manjunath SY, Venkataraman A. 2010. Rapid biosynthesis of irregular shaped gold nanoparticles from macerated aqueous extracellular dried clove buds (Syzygium aromaticum) solution. Colloids and Surfaces B: Biointerfaces 79, 235–240.

Dubey SP, Lahtinen M, Sillanpaa M. 2010. Green synthesis and characterizations of silver and gold nanoparticles using leaf extract of Rosa rugosa. Colloids and Surfaces A: Physicochem. Eng. Aspects 364, 34–41.

Gardea-Torresdey JL, Gomez E, Peralta-Videa JR, Parsons JG, Troiani H, Jose-Yacaman M. 2003. Alfalfa sprouts: A natural source for the synthesis of silver nanoparticles. Langmuir 19, 1357–1361.

Govindaraju K, Basha SK, Ganesh Kumar V, Singaravelu G. 2008. Silver, gold and bimetallic nanoparticles production using single-cell protein (Spirulina platensis Geitler). J Mater Sci 43, 5115–5122.

Govindaraju K, Tamilselvan S, Kiruthiga V, Singaravelu G. 2010. Biogenic silver nanoparticles by Solanum torvum and their promising antimicrobial activity. Journal of Biopesticides 3(1), 394–399.

Gurunathan S, Raman J, AbdMalek SN, John PA, Vikineswary S. 2013. Green synthesis of silver nanoparticles using Ganoderma neojaponicum Imazeki: a potential cytotoxic agent against breast cancer cells. Int. J. Nanomedicine 8, 4399-4413.

Kamat PV, Flumiani M, Hartland GV. 1998. Picosecond dynamics of silver nanoclusters: photo ejection of electrons and fragmentation. J. Phys. Chem. B 102, 3123–3128.

Khan Pathan R, Gali PR, Pathan P, Gowtham T, Pasupuleti S. 2012. In vitro antimicrobial activity of Citrus aurantifolia and its phytochemical screening. Asian Pac. J. Trop. Dis 2, S328–S331.

Klaus-Joerger T, Joerger R, Olsson E, Granqvist CG. 2001. Bacteria as workers in the living factory: metal-accumulating bacteria and their potential for materials science. Trends in Biotechnology 19(1), 15–20.

Kokila T, Ramesh PS, Geetha D. 2015. A biogenic approach for green synthesis of silver nanoparticles using peel extract of Citrus sinensis and its application. Int. J. Chem. Tech. Res 7(2), 804-813.

Kowshik M, Deshmukh N, Vogel W, Urban J, Kulkarni SK, Paknikar KM. 2002. Microbial synthesis of semiconductor CdS nanoparticles, their characterization, and their use in the fabrication of an ideal diode. Biotechnology and Bioengineering 78(5), 583–588.

Kumar KP, Paul W, Sharma CP. 2012. Green synthesis of silver nanoparticles with Zingiber officinale extract and study of its blood compatibility. BioNanoSci 2, 144–152.

Kumar R, Roopan SM, Prabhakarn A, Khanna VG, Chakroborty S. 2012. Agricultural waste Annona squamosa peel extract: biosynthesis of silver nanoparticles. Spectrochim. Acta Part A 90, 173–176.

Leela A, Vivekanandan M. 2008. Tapping the unexploited plant resources for the synthesis of silver nanoparticles. African Journal of Biotechnology 7(17), 3162–3165.

Mondal SBR, Mirdha S, Mahapatra C. 2009. The science behind sacredness of Tulsi (Ocimum sanctum Linn.). Indian Journal of Physiology and Pharmacology 53(4), 291–306.

Mustapha T, Ithnin NR, Othman H, Abu Hasan ZI, Misni N. 2023. Bio-fabrication of silver nanoparticles using Citrus aurantifolia fruit peel extract (CAFPE) and the role of plant extract in the synthesis. Plants 12(8), 1648.

Mustapha T, Misni N, Ithnin NR, Daskum AM, Unyah NZ. 2022. A review on plants and microorganisms mediated synthesis of silver nanoparticles, role of plant metabolites and applications. Int. J. Environ. Res. Public Health 19, 674.

Nur S, Othman A, Hassan MA, Nahar L, Basar N, Jamil S, Sarker SD. 2016. Essential oils from the Malaysian Citrus (Rutaceae) medicinal plants. Medicines 3, 13.

Pirtarighat S, Ghannadnia M, Baghshahi S. 2019. Green synthesis of silver nanoparticles using the plant extract of Salvia spinosa grown in vitro and their antibacterial activity assessment. J. Nanostructure Chem 9, 1–9.

Rafique M, Sadaf I, Rafique MS, Tahir MB. 2017. A review on green synthesis of silver nanoparticles and their applications. Artif. Cells Nanomed. Biotechnol 45, 1272–1291.

Raghunandan D, Borgaonkar PA, Bendegumble B, Bedre MD, Bhagawanraju M, Yalagatti MS, Huh DS, Abbaraju V. 2011. Microwave-assisted rapid extracellular biosynthesis of silver nanoparticles using carom seed (Trachyspermum copticum) extract and in vitro studies. American Journal of Analytical Chemistry 2, 475-483.

Rai M, Yadav A, Gade A. 2009. Silver nanoparticles as a new generation of antimicrobials. Biotechnol. Adv 27, 76–83.

Roy N, Barik A. 2010. Green synthesis of silver nanoparticles from the unexploited weed resources. International Journal of Nanotechnology 4, 95.

Sharma VK, Yingard RA, Lin Y. 2009. Silver nanoparticles: Green synthesis and their antimicrobial activities. Adv in Colloid and Interf Sci 145, 83-96.

Sujatha S, Tamilselvi Subha K, Panneerselvam1 A. 2013. Studies on biosynthesis of silver nanoparticles using mushroom and its antibacterial activities. Int. J. Curr. Microbiol. App. Sci 2(12), 605-614.

Source : Citrus aurantium bark,seeds, and leaves were used to synthesize and characterize silver nanoparticleand their antimicrobial activity was evaluated 

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