Home About us Editorial board Current issue Ahead of print Archives Submit article Instructions Subscribe Login  Contact Search

 Table of Contents  
Year : 2016  |  Volume : 7  |  Issue : 2  |  Page : 96-106

Significance of molecular markers in pharmacognosy: A modern tool for authentication of herbal drugs

1 Department of Pharmacy, Banasthali University, Tonk, Rajasthan, India
2 Department of Pharmacognosy and Phytochemistry, Faculty of Pharmacy, Jamia Hamdard, New Delhi, India

Date of Web Publication27-Sep-2016

Correspondence Address:
Sayeed Ahmad
Department of Pharmacognosy and Phytochemistry, Faculty of Pharmacy, Jamia Hamdard, New Delhi - 110 062
Login to access the Email id

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/2394-6555.191164

Rights and Permissions

Quality evaluation of herbal preparation is an elementary requirement of industry and other association dealing with Ayurvedic and herbal products. The growing use of botanical products now a days is forcing to assess these agents and to develop standards of quality and produce. An attempt has been made through this article to highlight the use of molecular markers for botanicals with special reference to Indian herbal medicine. As the desire for herbal-based products becomes ingrained in our society but standardization of botanicals offers many obstacles like the controversial identity of various plants, deliberated adulteration of plant material, ensuring quality is much more than discovery, specification, and process control. It also includes awareness of every aspect of a manufacturing process from research to shipping. Extensive research on DNA-based molecular markers is in progress for its great utility in the herbal drug analysis and widely used for the authentification of plant species of medicinal importance. DNA markers are reliable for information as the genetic composition is unique for each species and is not affected by age, physiological conditions, as well as environmental factors. DNA markers offer several advantages over conventional phenotypic markers, as they provide data that can be analyzed objectively.

Keywords: Botanicals, genetic markers, herbal drugs, quality control

How to cite this article:
Chester K, Tamboli ET, Paliwal SK, Ahmad S. Significance of molecular markers in pharmacognosy: A modern tool for authentication of herbal drugs. Drug Dev Ther 2016;7:96-106

How to cite this URL:
Chester K, Tamboli ET, Paliwal SK, Ahmad S. Significance of molecular markers in pharmacognosy: A modern tool for authentication of herbal drugs. Drug Dev Ther [serial online] 2016 [cited 2020 Jan 22];7:96-106. Available from: http://www.ddtjournal.org/text.asp?2016/7/2/96/191164

  Introduction Top

In the last two decades, DNA marker technologies have been revolutionized the plant pathogen genomic analysis and have been extensively employed in many fields of molecular plant pathology. Molecular markers offer also the possibility of faster and accurate identification and early detection of plant pathogen. [1],[2] DNA-based molecular markers have utility in the fields such as taxonomy, physiology, embryology, and genetics DNA-based techniques have been widely used for authentication of plant species of medicinal importance. [3]

  Molecular Marker Top

DNA sequence with a known location on a chromosome and associated with a particular gene or trait is known as a genetic marker. It is a variation, which may arise due to mutation or alteration in the genomic loci. A short DNA sequence may be a genetic marker, for example, a sequence surrounding a single base-pair change (single nucleotide polymorphism), or like mini satellites - A long one [Figure 1]. [4]
Figure 1: Desirable properties of ideal DNA markers

Click here to view

Desirable properties of ideal DNA markers

  • Easily available
  • Assay is easy and rapid
  • Highly polymorphic and reproducible
  • Co-dominant inheritance and recurrent occurrence in genome
  • Selectively neutral to environmental conditions or management practices
  • Data exchange between different laboratories should be easy
  • High genomic abundance.

It is really difficult to get a molecular marker of above criteria. Depending on the type of study undertaken, a marker system can be recognized that would fulfill the above characteristics. [5]

  Basic Molecular Marker Techniques Top

Basic marker techniques can be classified into three categories:

  1. Polymerase chain reaction (PCR)-based techniques
  2. Non-PCR-based techniques or hybridization based techniques
  3. Microsatellite-based marker techniques [Figure 2].
    Figure 2: Basic molecular marker techniques

    Click here to view

Polymerase chain reaction-based techniques

In the case of PCR-based markers, the primers of known sequence and length are used to amplify genomic and cDNA sequences which are visualized by gel electrophoresis technique. The invention of PCR which is a very versatile and extremely sensitive technique, [6] contributes to use a thermostable DNA polymerase and lead to the development of various molecular marker techniques. [7]

Randomly amplified polymorphic DNA markers

A single species of primer anneals to the genomic DNA at two different sites on complimentary strands of the DNA template. After PCR amplification, a discrete DNA product is obtained if these priming sites are within the amplification range of each other. The introduction of this system produces amplification of several discrete loci. [8]

DNA amplification fingerprinting

In this technique, a single arbitrary primer of only five bases is used to amplify the DNA by PCR. For this marker assay gives simple banding patterns, much-optimized reaction conditions are required and are useful for DNA fingerprinting. Such banding patterns are analyzed by polyacrylamide gel electrophoresis. [9] Arbitrary primed PCR (AP-PCR).

DNA amplification patterns are obtained using single primer of 10-50 bases long in PCR and annealing is carried out under nonstringent conditions. [10]

Sequence characterized amplified regions

Sequence characterized amplified regions (SCARs) are similar to sequence tagged sites (STS) markers but in comparison to random amplified polymorphic DNA (RAPD) they are more reproducible. Although SCARS are mostly dominant markers, also behave as co-dominant markers by digesting them with tetra cutting restriction enzymes. Sex identification of papaya has been carried out using SCAR marker. [11]

Cleaved amplification polymorphic sequence

PCR primers for this process can be synthesized based on sequence information in databank and the electrophoretic patterns are obtained using restriction enzyme digestion of the PCR products. [12],[13]

Randomly amplified microsatellite polymorphism

The methodology of these PCR based markers is that first the genomic DNA is amplified using the arbitrary (RAPD) primers. The amplified products thus obtained are then separated electrophoretically and the dried gel is hybridized with microsatellite oligonucleotide probes. Many advantages of oligonucleotide fingerprinting, [14] RAPD [15] and microsatellite primed-PCR are thus combined in randomly amplified microsatellite polymorphism. [16],[17] Advantages include speed of the assay, high sensitivity, high level of variability detected and no requirement of prior DNA sequence information. [18]

  Amplified Fragment Length Polymorphism Top

Amplified fragment length polymorphism (AFLP) was developed for detection of genomic restriction fragments by PCR amplification, thus fingerprinting patterns are obtained. AFLP is an ingenious combination of restriction fragment length polymorphism (RFLP) and PCR. [19] In the detection of polymorphism between closely related genotypes AFLP is extremely useful. [20],[21]

  Nonpolymerase Chain Reaction-based Techniques Top

In the hybridization-based markers or non-PCR based marker, the DNA profiles are visualized by hybridizing the restriction enzyme digested DNA to a labeled probe which is a DNA fragment of known/unknown sequence.

Restriction fragment length polymorphism

The RFLP analysis consists of, restriction endonuclease digested genomic DNA is resolved by gel electrophoresis and then blotted on to a nitrocellulase membrane. [22] Specific banding patterns are then visualized by hybridization with a labeled probe. RFLP are very reliable markers on linkage analysis and breeding and are co-dominant in nature. To constructs a genetic map RFLPs were used for the 1 st time to construct a genetic map. [23]

Detailed below are the modifications of the RFLP marker system.

Sequence tagged sites

In this, RFLP probes specifically linked to a desired trait can be converted into PCR based STS oligonucleotide primers based on nucleotide sequence of the probe giving a polymorphic band pattern. This is extremely useful for studying the relationship among several species at a specific locus. [24]

Allele specific associated primers

Specific allele is sequenced and on the base of this sequence, specific primers are designed for DNA template amplification. From this, a single fragment at stringent annealing temperature condition is obtained, and there are used to tag given plant and the gene of interest. [25]

Expressed sequence tag markers

These are introduced for obtaining partial sequencing of random cDNA clones. They are also useful in genome sequencing and mapping programs. [26]

Single strand conformation polymorphism

This is a powerful and popularly used technique for detection of point mutations. It can identify heterozygosity of DNA fragments of the same molecular weight. [27]

  Microsatellites and Minisatellites Top

In virtually all eukaryotic species, 30-90% of the genome is constituted of repetitive DNA, and its nature is highly polymorphic. Microsatellites and minisatellites is one major form of repetitive DNA. [28] With a monomer repeat length of about 11-60 bp, microsatellites are short tandem repeats or simple sequence repeats of 1-6 bp length, repeated several times. Thus, micro and minisatellites form an ideal marker system which simultaneously create complex banding system and detect multiple DNA loci simultaneously. These dominant fingerprinting markers, exhibit high level heterozygosity, and are of Mendelian inheritance. [29]

Detailed below are minisatellite and microsatellite sequence based markers.

Sequence tagged microsatellites sites

Using specific primers designed from sequence 22 data of a specific loci, DNA polymorphism is detected. Primers complementary to the flanking regions of the simple sequence repeat loci yield high polymorphism. For clear banding pattern Di-, tri- and tetra-nucleotide microsatellites are more popular for sequence tagged microsatellites sites analysis. [30] For diversity analysis, dinucleotides which are generally abundant in the genome have been used. [31]

Directed amplification of minisatellite-region DNA

In this case, minisatellites are used as primers for DNA amplification. It is introduced for the 1 st time and is found to be useful for species differentiation and cultivar identification. [32],[33]

Inter simple sequence repeat markers

This technique was reported for amplifying genomic DNA at the 3'end. They are mostly dominant markers. Number of primers can be synthesized for various combinations of di-, tri-, and tetra and penta-nucleotides. [34]

  Advances in Molecular Marker Techniques Top

Molecular marker techniques have made advances through incorporation of modification in the methodology leads to evolution of several basic techniques.

Organelle microsatellites

Chloroplast DNA and mitochondrial DNA are considered to study the genetic structure and phylogenetic relationships in plants, organelle and genome. Compared to nuclear alleles, chloroplast and mitochondrial genomes, due to their uniparental mode of transmission, exhibit different patterns of genetic differentiation. [35] Thus, for a widespread understanding of plant population delineation and evolution, three interrelated genomes must be considered, namely nuclear microsatellites, chloroplast and mitochondrial microsatellites have also been developed. Chloroplast microsatellites consisting of relatively short and several mononucleotide sequences are ubiquitous and polymorphic components of chloroplast DNA. [36] Chloroplast genome-based markers uncover genetic discontinuities and distinctiveness among or between taxa with slight morphological differentiation, which nuclear DNA markers cannot reveal sometime. [37]

Sequence-related amplified polymorphism

Sequence-related amplified polymorphism (SRAP) is based on two-primer amplification, mainly 17-21 nucleotides in length. For polymorphism detection SCRAP uses pairs of primers with AT- or GC-rich cores to amplify intragenic fragments. The amplification of open reading frames is the aim of SCRAP technique, SRAP combines simplicity, reliability, moderate throughput ratio, and facile sequencing of selected bands. [38]

Target region amplification polymorphism

The Target region amplification polymorphism technique is a PCR-based convenient technique, which utilizes expressed sequence tag (EST) database information and bioinformatics to generate around targeted candidate gene sequences, polymorphic markers. The fixed primer is designed from the targeted EST sequence in the database; the second primer is an arbitrary primer with either an AT- or GC-rich core to anneal with an intron or exon. [39]

Transposable elements-based molecular markers

The mobile genetic elements which are capable of changing their location in the genome are known as transposons and were discovered in maize almost 60 years ago. Transposable elements, consists of two broad classes and each with its own characteristics properties. [40] Retroelements, such as retrotransposons, short interspersed nuclear elements, and long interspersed nuclear elements, it is the element-encoded mRNA, and not the element itself, that forms the transposition intermediate. The original copy remains intact at the donor site each transposition event creates a new copy of transposons. In contrast, Class II consists of DNA transposons, which change their location in the genome by a "cut and paste" mechanism.[41]

Retrotransposon-based molecular markers

Retrotransposons are the major class of repetitive DNA comprising 40-60% of the entire genome in plants with large genomes. [42] Retrotransposons can be divided into three categories based on structural organization and amino acid similarities among their encoded reverse transcriptases. Long terminal direct repeats (LTRs) flank two of these categories and they encode proteins similar to the retroviruses. These LTR-retrotransposons are known as gypsy-like and copia-like retrotransposons. The LINE1-like or non-LTR retrotransposons are third class of retrotransposons and they lack terminal repeats and encode proteins with significantly less similarity to those of the retroviruses. The replicating process of retrotransposons is by successive transcription, reverse transcription, and insertion of the new cDNA copies back into the genome, copia-like [43],[44] and gypsy-like retrotransposons [45] are present throughout the plant kingdom.

Inter-retrotransposon amplified polymorphism and retrotransposon-microsatellite amplified polymorphism

Based on the position of given LTRs within the genome inter-retrotransposon amplified polymorphism (IRAP) and REMP are two amplification based marker methods which have been developed. Proximity of two LTRs using outward-facing primers annealing to LTR target sequences generate the IRAP markers. In retrotransposon-microsatellite amplified polymorphism, amplification between LTRs proximal to simple sequence repeats such as constitutive microsatellites produces markers. [45]

Sequence-specific amplification polymorphism

The technique was first used to investigate the location of BARE-1 retrotransposons in the barley genome. [46] In principle, it is a simple modification of the standard AFLP protocol. [47]

Retrotransposon-based insertion polymorphism

Using the PDR1 retrotransposon in the pea, this technique was developed this technique requires the sequence information of the 50 and 30 regions flanking the transposons. [48]

  RNA-based Molecular Markers Top

Biological responses and developmental programming are regulated by the precise control of genetic expression. Obtaining in depth information about these processes necessitates the study of differential patterns of gene expression. PCR-based marker techniques, such as, cDNA, AFLP, and RNA fingerprinting by arbitrarily primed Polymerase chain reaction (RAP-PCR) are used for differential RNA study selective amplification of cDNAs. Replicated tests show that cDNA-single strand conformation polymorphism reliably separates duplicated transcripts with 99% sequence identity. [49]

RNA fingerprinting by arbitrarily primed polymerase chain reaction

Arbitrarily primer at low stringency for first and second strand cDNA synthesis followed by PCR amplification of cDNA population involves fingerprinting of RNA populations. The method requires nanograms of total RNA and is unaffected by low levels of genomic DNA contamination. [50]

cDNA-amplified fragment length polymorphism

A novel RNA fingerprinting technique to display differentially expressed genes is cDNA-AFLP technique. [51] The methodology includes digestion of cDNAs by two restriction enzymes followed by ligation of oligonucleotide adapters and PCR amplification using primers complementary to the adapter sequences with additional selective nucleotides at the 30 end. [52] The cDNA-AFLP technique is a more stringent and reproducible than RAP-PCR. [53]

  DNA Barcoding Top

Apart from other technique, it is a dominant method for species identification and discovery since other methods have various limitations and cannot be used in a large scale or in an efficient manner. DNA barcoding uses a short genetic marker in an organism's DNA to identify it as belonging to a particular species. It distinguishes from molecular phylogeny in that the main aim is not to determine classification but to identify an unknown sample in terms of a known classification. [54] Although barcodes are sometimes used in an effort to identify unknown species or assess whether species should be combined or separated, such usage, if possible at all, pushes the limits of what are barcodes capabilities. [55]

Comparison of various aspects of widely used molecular marker techniques [Table 1].
Table 1: Examples of some medicinal plants and different marker used for various studies and their results

Click here to view

[TAG:2]Applications are Prospect of Pharmaceutical Sciences[/TAG:2]

Pharmacognostic application

Pharmacognosy generally related to quality-control issues using routine organoleptic parameters of crude drugs. DNA-based techniques have been widely used for authentication of plant species of medicinal importance. Since certain rare and expensive medicinal plant species are often adulterated or substituted by morphologically similar, easily available, or less expensive species. For example, Swertia chirata is frequently adulterated or substituted by the cheaper Andrographis paniculata, therefore, additional methods of identification at the species level have been sought and genome-based methods have been developed for the identification of medicinal plants starting in the early 1990's. [56],[57],[58],[59] This work was greatly facilitated by the invention of the PCR and the introduction of a heat-stable DNA polymerase from the thermophilic bacterium. At present, a practical and powerful tool, i.e. DNA barcodes, is developed for identifying medicinal plants and their adulterants in trade and for ensuring safety in their use. [60] (CBOL Plant Working Group, 2009). Among the PCR-based molecular techniques, RAPD is convenient in performance and does not require any information about the DNA sequence to be amplified. [61] Due to its procedural simplicity, the use of RAPD as molecular markers for taxonomic and systematic analyses of plants. [62] As well as in plant breeding and the study of genetic relationships, has considerably increased. [63] Recently, RAPD has been used for the estimation of genetic diversity in various endangered plant species. [64],[65],[66],[67],[68]

This technique remains important in plant genome research with its applications in pharmacognostic identification and analysis.

Pharmacological application

Genetic markers can be used to study the relationship between an inherited disease and its genetic cause (e.g., a particular mutation of a gene that results in a defective protein). Genetic markers have to be easily identifiable, associated with a specific locus, and highly polymorphic, because homozygotes do not provide any information. The methods used to study the genome or phylogenetics is RFLP, AFLP, RAPD, SSR. They can be used to create genetic maps of whatever organism is being studied. The presence of different alleles due to a distorted segregation at the genetic markers is indicative of the difference between selected and nonselected livestock. Endosomal sorting receptors, useful as molecular markers to define plant endosomal compartments are, for example, the vacuolar sorting receptor (VSR) BP-80 from pea (Pisum sativum) and its Arabidopsis homolog AtELP1, which are known to predominantly associate with prevacuolar compartments/multivesicular bodies. [68]

Pharmaceutical application

Molecular markers in pharmaceutics relate to drugs and drug development, for example, in silico drug profiling of the human kinome based on a molecular marker for cross-reactivity. [69] Molecular markers focus on molecular mechanistic approaches to the development of bio-available drugs as well as concentrates on the integration of applications of the chemical and biological sciences to advance the development of new drugs and delivery systems.

Other application in plant genome

Examples of some medicinal plants and different marker used for various studies [Table 2].
Table 2: Comparison of various aspects of widely used molecular marker techniques

Click here to view

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

  References Top

Bridge PD, Singh T, Arora DK. The application of molecular markers in the epidemiology of plant pathogenic fungi. In: Arora DK, Bridge PD, Bhatnagar D, editors. Fungal Biotechnology in Agricultural, Food, and Environmental Applications. USA: CRC Press; 2003.  Back to cited text no. 1
Schmidt H, Taniwaki MH, Vogel RF, Niessen L. Utilization of AFLP markers for PCR-based identification of Aspergillus carbonarius and indication of its presence in green coffee samples. J Appl Microbiol 2004;97:899-909.  Back to cited text no. 2
Joshi K, Chavan P, Warude D, Patwardhan B. Molecular markers in herbal drug technology. Curr Sci India 2004;87:2.  Back to cited text no. 3
Srivastava S, Mishra N. Genetic markers - A cutting-edge technology in herbal drug research. J Chem Pharm Res 2009;1:1-18.  Back to cited text no. 4
Saiki RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, et al. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 1985;230:1350-4.  Back to cited text no. 5
Saiki RK, Gelfand DH, Stoffei S, Scharf SJ, Higuchi R, Horn GT, et al. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Nature 1988;239:487-97.  Back to cited text no. 6
Welsh J, McClelland M. Fingerprinting genomes using PCR with arbitrary primers. Nucleic Acids Res 1990;18:7213-8.  Back to cited text no. 7
Caetano-Anollés G, Bassam BJ, Gresshoff PM. DNA amplification fingerprinting using very short arbitrary oligonucleotide primers. Biotechnology (N Y) 1991;9:553-7.  Back to cited text no. 8
Welsh J, Honeycutt RJ, McClelland M, Sobral BW. Parentage determination in maize hybrids using the arbitrarily primed polymerase chain reaction (AP-PCR). Theor Appl Genet 1991;82:473-6.  Back to cited text no. 9
Parasnis AS, Gupta VS, Tamhankar SA, Ranjekar PK. A highly reliable sex diagnostic PCR assay for mass screening of papaya seedlings. Mol Breed 2000;6:337-44.  Back to cited text no. 10
Konieczny A, Ausubel FM. A procedure for mapping Arabidopsis mutations using co-dominant ecotype-specific PCR-based markers. Plant J 1993;4:403-10.  Back to cited text no. 11
Jarvis P, Lister C, Szabo V, Dean C. Integration of CAPs markers into the RFLP map generated using recombinant inbred lines of Arabidopsis thaliana. Plant Mol Biol 1994;24:685-7.  Back to cited text no. 12
Epplen JT. The methodology of multilocus DNA fingerprinting using radioactive or nonradioactive oligonucleotide probe specific for simple repeat motifs. Adv Electrophor 1992;5:55-112.  Back to cited text no. 13
Williams JG, Kubelik AR, Livak KJ, Rafalski JA, Tingey SV. DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic Acids Res 1990;18:6531-5.  Back to cited text no. 14
Weising K, Nybom H, Wolff K, Meyer W. DNA fingerprinting of plants and fungi. USA: Boca Raton. CRC Press; 1995. p. 1-3.  Back to cited text no. 15
Gupta M, Chyi YS, Romero-Severson J, Owen JL. Amplification of DNA markers from evolutionarily diverse genomes using single primers of simple-sequence repeats. Theor Appl Genet 1994;89:998-1006.  Back to cited text no. 16
Richardson T, Cato S, Ramser J, Kahl G, Weising K. Hybridization of microsatellites to RAPD: A new source of polymorphic markers. Nucleic Acids Res 1995;23:3798-9.  Back to cited text no. 17
Zabeau M. Selective Restriction Fragment Amplification a General Method for DNA Fingerprinting. European Patent Application Publication No. 0534858A1; 1993.  Back to cited text no. 18
Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, et al. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science 1988;239:487-91.  Back to cited text no. 19
Erlich HA, Gelfand D, Sninsky JJ. Recent advances in the polymerase chain reaction. Science 1991;252:1643-51.  Back to cited text no. 20
Southern EM. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 1975;98:503-17.  Back to cited text no. 21
Botstein D, White RL, Skolnick M, Davis RW. Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am J Hum Genet 1980;32:314-31.  Back to cited text no. 22
Bustos A, Solier C, Jouve N. Analysis of PCR-based markers using between species of Hordeum (Poaceae). Genome 1999;42:129-38.  Back to cited text no. 23
Gu WK, Weeden NF, Yu J, Wallace DH. Large-scale, cost-effective screening of PCR products in marker-assisted selection applications. Theor Appl Genet 1995;91:465-70.  Back to cited text no. 24
Adams MD, Kelley JM, Gocayne JD, Dubnick M, Polymeropoulos MH, Xiao H, et al. Complementary DNA sequencing: Expressed sequence tags and human genome project. Science 1991;252:1651-6.  Back to cited text no. 25
Orita M, Suzuki Y, Sekiya T, Hayashi K. Rapid and sensitive detection of point mutations and DNA polymorphisms using the polymerase chain reaction. Genomics 1989;5:874-9.  Back to cited text no. 26
Litt M, Luty JA. A hypervariable microsatellite revealed by in vitro amplification of a dinucleotide repeat within the cardiac muscle actin gene. Am J Hum Genet 1989;44:397-401.  Back to cited text no. 27
Jeffreys AJ, Wilson V, Thein SL. Hypervariable 'minisatellite' regions in human DNA. Nature 1985;314:67-73.  Back to cited text no. 28
Hearne CM, Ghosh S, Todd JA. Microsatellites for linkage analysis of genetic traits. Trends Genet 1992;8:288-94.  Back to cited text no. 29
Rafalski JA, Vogel JM, Morgante M, Powell W, Andre C, Tingey SV. Generating and using DNA markers in plants. In: Birren B, Lai E, editors. Nonmammalian Genomic Analysis. San Diego, CA, USA: Academic Press; 1996. p. 75-134.  Back to cited text no. 30
Heath DD, Iwama GK, Devlin RH. PCR primed with VNTR core sequences yields species specific patterns and hypervariable probes. Nucleic Acids Res 1993;21:5782-5.  Back to cited text no. 31
Somers DJ, Zhou Z, Bebeli PJ, Gustafson JP. Repetitive, genome-specific probes in wheat (Triticum aestivum L. em Thell) amplified with minisatellite core sequences. Theor Appl Genet 1996;93:982-9.  Back to cited text no. 32
Zietkiewicz E, Rafalski A, Labuda D. Genome fingerprinting by simple sequence repeat (SSR)-anchored polymerase chain reaction amplification. Genomics 1994;20:176-83.  Back to cited text no. 33
Provan J, Russell JR, Booth A, Powell W. Polymorphic chloroplast simple sequence repeat primers for systematic and population studies in the genus Hordeum Mol Ecol 1999;8:505-11.  Back to cited text no. 34
Powell W, Morgante M, McDevitt R, Vendramin GG, Rafalski JA. Polymorphic simple sequence repeat regions in chloroplast genomes: Applications to the population genetics of pines. Proc Natl Acad Sci U S A 1995;92:7759-63.  Back to cited text no. 35
Wolfe KH, Li WH, Sharp PM. Rates of nucleotide substitution vary greatly among plant mitochondrial, chloroplast, and nuclear DNAs. Proc Natl Acad Sci U S A 1987;84:9054-8.  Back to cited text no. 36
Provan J, Powell W, Hollingsworth PM. Chloroplast microsatellites: New tools for studies in plant ecology and evolution. Trends Ecol Evol 2001;16:142-7.  Back to cited text no. 37
Chung SM, Staub JE. The development and evaluation of consensus chloroplast primer pairs that possess highly variable sequence regions in a diverse array of plant taxa. Theor Appl Genet 2003;107:757-67.  Back to cited text no. 38
Li G, Quiros CF. Sequence-related amplified polymorphism (SRAP), a new marker system based on a simple PCR reaction: Its application to mapping and gene tagging in Brassica. Theor Appl Genet 2001;103:455-546.  Back to cited text no. 39
Finnegan DJ. Eukaryotic transposable elements and genome evolution. Trends Genet 1989;5:103-7.  Back to cited text no. 40
Grzebelus D. Transposon insertion polymorphism as a new source of molecular markers. J Fruit Ornamental Plant Res 2006;14:21-9.  Back to cited text no. 41
Kumar A, Bennetzen JL. Plant retrotransposons. Annu Rev Genet 1999;33:479-532.  Back to cited text no. 42
Voytas DF, Cummings MP, Koniczny A, Ausubel FM, Rodermel SR. copia-like retrotransposons are ubiquitous among plants. Proc Natl Acad Sci U S A 1992;89:7124-8.  Back to cited text no. 43
Kumar A. The adventures of the Ty1-copia group of retrotransposons in plants. Trends Genet 1996;12:41-3.  Back to cited text no. 44
Kalendar R, Grob T, Regina M, Suoniemi A, Schulman A. IRAP and REMAP: Two new retrotransposon-based DNA fingerprinting techniques. Theor Appl Genet 1999;98:704-11.  Back to cited text no. 45
Waugh R, McLean K, Flavell AJ, Pearce SR, Kumar A, Thomas BB, et al. Genetic distribution of Bare-1-like retrotransposable elements in the barley genome revealed by sequence-specific amplification polymorphisms (S-SAP). Mol Gen Genet 1997;253:687-94.  Back to cited text no. 46
Vos P, Hogers R, Bleeker M, Reijans M, van de Lee T, Hornes M, et al. AFLP: A new technique for DNA fingerprinting. Nucleic Acids Res 1995;23:4407-14.  Back to cited text no. 47
Flavell AJ, Knox MR, Pearce SR, Ellis TH. Retrotransposon-based insertion polymorphisms (RBIP) for high throughput marker analysis. Plant J 1998;16:643-50.  Back to cited text no. 48
Cronn RC, Adams KL. Quantitative analysis of transcript accumulation from genes duplicated by polyploidy using cDNA-SSCP. Biotechniques 2003;34:726-30, 732, 734.  Back to cited text no. 49
Welsh J, Chada K, Dalal SS, Cheng R, Ralph D, McClelland M. Arbitrarily primed PCR fingerprinting of RNA. Nucleic Acids Res 1992;20:4965-70.  Back to cited text no. 50
Bachem CW, van der Hoeven RS, de Bruijn SM, Vreugdenhil D, Zabeau M, Visser RG. Visualization of differential gene expression using a novel method of RNA fingerprinting based on AFLP: Analysis of gene expression during potato tuber development. Plant J 1996;9:745-53.  Back to cited text no. 51
Bachem CW, Oomen RJ, Visser GF. Transcript imaging with cDNA-AFLP: A step-by-step protocol. Plant Mol Biol Rep 1998;16:157.  Back to cited text no. 52
Liang P, Pardee AB. Differential display of eukaryotic messenger RNA by means of the polymerase chain reaction. Science 1992;257:967-71.  Back to cited text no. 53
Kress WJ, Wurdack KJ, Zimmer EA, Weigt LA, Janzen DH. Use of DNA barcodes to identify flowering plants. Proc Natl Acad Sci U S A 2005;102:8369-74.  Back to cited text no. 54
Seberg O, Petersen G. How many loci does it take to DNA barcode a crocus? PLoS One 2009;4:e4598.  Back to cited text no. 55
Shaw PC, Wang J, But PP. Authentication of Chinese Medicinal Materials by DNA Technology. Singapore: World Science; 2002.  Back to cited text no. 56
Techen N, Crockett SL, Khan IA, Scheffler BE. Authentication of medicinal plants using molecular biology techniques to compliment conventional methods. Curr Med Chem 2004;11:1391-401.  Back to cited text no. 57
Sucher NJ, Carles MC. Genome-based approaches to the authentication of medicinal plants. Planta Med 2008;74:603-23.  Back to cited text no. 58
Hao da C, Yang L, Huang B. Molecular evolution of paclitaxel biosynthetic genes TS and DBAT of Taxus species. Genetica 2009;135:123-35.  Back to cited text no. 59
Chen S, Yao H, Han J, Liu C, Song J, Shi L, et al. Validation of the ITS2 region as a novel DNA barcode for identifying medicinal plant species. PLoS One 2010;5:e8613.  Back to cited text no. 60
Weder JK. Influence of experimental conditions on the reproducibility of RAPD-PCR identification of legumes and cereals. Lebensm Wiss Technol 2002;35:233-8.  Back to cited text no. 61
Bartish IV, Garkava LP, Rumpunen K, Nybom H. Phylogenetic relationship and differentiation among and within population of Chaenomeles Lindl. (Rosaceae) estimated with RAPDs and isozyme. Theor Appl Genet 2000;101:554-63.  Back to cited text no. 62
Ranade SA, Farooqui N, Bhattacharya E, Verma A. Gene tagging with random amplified polymorphic DNA (RAPD) marker for molecular breeding in plants. Crit Rev Plant Sci 2001;20:251-75.  Back to cited text no. 63
Wang C, Zhou TH, Yang X, Guo J, Zhao GF. Identification of seven plants of Gynostemma BL. by ISSR-PCR. Chin Tradit Herbal Drugs 2008;39:588-91.  Back to cited text no. 64
Lu HP, Cai YW, Chen XY, Zhang X, Gu YJ, Zhang GF. High RAPD but no cpDNA sequence variation in the endemic and endangered plant, Heptacodium miconioides Rehd. (Caprifoliaceae). Genetica 2006;128:409-17.  Back to cited text no. 65
Liu P, Yang YS, Hao CY, Guo WD. Ecological risk assessment using RAPD and distribution pattern of a rare and endangered species. Chemosphere 2007;68:1497-505.  Back to cited text no. 66
Li G, Quiros CF. Sequence-related amplified polymorphism (SRAP), a new marker system based on a simple PCR reaction: Its application to mapping and gene tagging in Brassica. Theor Appl Genet 2001;103:455-546.  Back to cited text no. 67
Tse YC, Mo B, Hillmer S, Zhao M, Lo SW, Robinson DG, et al. Identification of multivesicular bodies as prevacuolar compartments in Nicotiana tabacum BY-2 cells. Plant Cell 2004;16:672-93.  Back to cited text no. 68
Zhang X, Fernández A. In silico drug profiling of the human kinome based on a molecular marker for cross reactivity. Mol Pharm 2008;5:728-38.  Back to cited text no. 69
Mehdi RM, Tabatabaei BE, Arzani A, Etemad N. Assessment of genetic diversity among and within Achillea species using amplified fragment length polymorphism (AFLP). Biochem Syst Ecol 2009;37:354-61.  Back to cited text no. 70
Ahlawat A, Katoch M, Ram G, Ahuja A. Genetic diversity in Acorus calamus L. as revealed by RAPD markers and its relationship with β-asarone content and ploidy level. Sci Hortic 2010;124:294-7.  Back to cited text no. 71
Ge XJ, Yu Y, Yuan YM, Huang HW, Yan C. Genetic diversity and geographic differentiation in endangered Ammopiptanthus (Leguminosae) populations in desert regions of Northwest China as revealed by ISSR analysis. Ann Bot 2005;95:843-51.  Back to cited text no. 72
Lin SF, Tsay HS, Chou TW, Yang MJ, Cheng KT. Genetic variation of Anoectochilus formosanus revealed by ISSR and AFLP analysis. J Food Drug Anal 2007;15:156-62.  Back to cited text no. 73
Singh A, Chaudhury A, Srivastava PS, Lakshmikumaran M. Comparison of AFLP and SAMPL markers for assessment of intra-population genetic variation in Azadirachta indica A. Juss. Plant Sci 2002;162:17-25.  Back to cited text no. 74
Thendral BH, Premalakshmi V, Sekar T. Genetic diversity in Azima tetracantha (Lam) assessed through RAPD analysis. Indian J Sci Technol 2010;3:685-7.  Back to cited text no. 75
Darokar MP, Khanuja SP, Shasany AK, Kumar S. Low levels of genetic diversity detected by RAPD analysis in geographically distinct accessions of Bacopa monnieri. Genet Resour Crop Evol 2001;48:555-8.  Back to cited text no. 76
Vanijajiva O, Sirirugsa P, Suvachittanont W. Confirmation of relationships among Boesenbergia (Zingiberaceae) and related genera by RAPD. Biochem Syst Ecol 2005;33:159-70.  Back to cited text no. 77
Shaw RK, Charya L, Mukherjee AK. Assessment of genetic diversity in a highly valuable medicinal plant Catharanthus roseus using molecular markers. Crop Breed Appl Biotechnol Braz Soc Plant Breed 2009;9:52-9.  Back to cited text no. 78
Qiu YX, Hong DY, Fu CX, Cameron KM. Genetic variation in the endangered and endemic species Changium smyrnioides (Apiaceae). Biochem Syst Ecol 2004;32:583-96.  Back to cited text no. 79
Shao QS, Guo QS, Deng YM, Guo HP. A comparative analysis of genetic diversity in medicinal Chrysanthemum morifolium based on morphology, ISSR and SRAP marker. Biochem Syst Ecol 2010;38:1160-9.  Back to cited text no. 80
Djè Y, Tahi GC, Zoro Bi IA, Malice M, Baudoin JP, Bertin P. Optimization of ISSR marker for African edible-seeded Cucurbitaceae species' genetic diversity analysis. Afr J Biotechnol 2006;5:83-7.  Back to cited text no. 81
Mandal AB, Thomas VA, Elanchezhian R. RAPD pattern of Costus specious Koen ex. Retz., an important medicinal plant from the Andaman and Nicobar Islands. Port Blair, India: Biotechnology Section, Central Agricultural Research Institute; 2007.  Back to cited text no. 82
Paris HS, Yonash N, Portnoy V, Mozes-Daube N, Tzuri G, Katzir N. Assessment of genetic relationships in Cucurbita pepo (Cucurbitaceae) using DNA markers. Theor Appl Genet 2003;106:971-8.  Back to cited text no. 83
Tyagi RK, Agrawal A, Mahalakshmi C, Hussain Z, Tyagi H. Low-cost media for in vitro conservation of turmeric (Curcuma longa L.) and genetic stability assessment using RAPD markers. In Vitro Cell Dev Biol Plant 2007;43:51-8.  Back to cited text no. 84
Khan S, Mirza KJ, Abdin MZ. Development of RAPD markers for authentication of medicinal plant Cuscuta reflexa. Eur J Bio Sci 2007;4:1-7.  Back to cited text no. 85
Ding G, Li X, Ding X, Qian L. Genetic diversity across natural populations of Dendrobium officinale, the endangered medicinal herb endemic to China revealed by ISSR and RAPD markers. Russ Genet 2009;45:375-82.  Back to cited text no. 86
Gavidiai I, Agudo LD, Bermúdez PP. Digitalis obscura selection and long-term cultures of high-yielding Digitalis obscura plants RAPD markers for analysis of genetic stability. Sci Hortic 1996;121:197-205.  Back to cited text no. 87
Chuang SJ, Chen CL, Chen JJ, Wei YC, Sung JM. Detection of somaclonal variation in micro-propagated Echinacea purpurea using AFLP marker. Sci Hortic 2009;120:121-6.  Back to cited text no. 88
Prakash S, Grobbelaar N, Van Staden J. Diversity in Encephalartos woodii collections based on Random Amplified DNA markers (RAPD's) and Inter-Specific Sequence Repeats (ISSR's). S Afr J Bot 2008;74:341-4.  Back to cited text no. 89
Birmeta G, Nybom H, Bekele E. RAPD analysis of genetic diversity among clones of the Ethiopian crop plant Ensete ventricosum Euphytica. J Genet 2002;124:315-25.  Back to cited text no. 90
Guasmi F, Ferchichi A, Farés K, Touil L. Identification and differentiation of Ficus carica L. cultivars using inter simple sequence repeat markers. Afr J Biotechnol 2006;5:1370-4.  Back to cited text no. 91
Albani MC, Battey NH, Wilkinson MJ. The development of ISSR-derived SCAR markers around the Seasonal Flowering Locus (SFL) in Fragaria vesca. Theor Appl Genet 2004;109:571-9.  Back to cited text no. 92
Xiong H, Luo B, Zheng Y, Cui Y, Zhang Y, Wu Q, et al. Genomic DNA extraction and RAPD analysis for Gastrodia elata Bl. (Orchidaceae) using an improved method. J Bio Sci 2012;12:206-8.  Back to cited text no. 93
Kaur R, Panwar N, Saxena B, Raina R, Bharadwaj SV. Genetic stability in long-term micropropagated plants of Gentiana Kurroo - An endangered medicinal plant. J New Seed 2009;10:236-244.  Back to cited text no. 94
Fracaro F, Echeverrigaray S. Genetic variability in Hesperozygis ringens Benth. (Lamiaceae), an endangered aromatic and medicinal plant of southern Brazil. Biochem Genet 2006;44:471-82.  Back to cited text no. 95
Yong ZY, Geng Y, Tashi TT, Liu N, Wang Q, Zhong Y. High genetic differentiation and low genetic diversity in Incarvillea younghusbandii, an endemic, plants of Qinghai-Tibetan Plateau, revealed by AFLP markers. Biochem Syst Ecol 2009;37:589-96.  Back to cited text no. 96
Devaiah K, Balasubramani SP, Venkatasubramanian P. Development of randomly amplified polymorphic DNA based SCAR marker for identification of Ipomoea mauritiana Jacq (Convolvulaceae). Evid Based Complement Alternat Med 2011;2011:868720.  Back to cited text no. 97
Basha SD, Sujatha M. Genetic analysis of Jatropha species and interspecific hybrid of Jatropha curcas using nuclear and organelle specific markers. Euphytica 2009;168:197-214.  Back to cited text no. 98
Ci XQ, Chen JQ, Li QM, Li J. AFLP and ISSR analysis reveals high genetic variation and inter-population differentiation in fragmented populations of the endangered Litsea szemaois (Lauraceae) from Southwest China. Plant Syst 2008;273:237-46.  Back to cited text no. 99
Talhinhas PP, Leita J, Neves-Martins J. Collection of Lupinus angustifolius L. germplasm and characterization of morphological and molecular diversity. Genet Resour Crop Evol 2006;53:563-78.  Back to cited text no. 100
Yu HH, Yang ZL, Sun B, Liu RN. Genetic diversity and relationship of endangered plant Magnolia officinalis (Magnoliaceae) assessed with ISSR polymorphism. Biochem Syst Ecol 2011;39:71-8.  Back to cited text no. 101
Solouki M, Mehdikhani H, Zeinali H, Emamjomeh AA. Study of genetic diversity in Chamomile (Matricaria chamomilla) based on morphological traits and molecular marker. Sci Hortic 2008;117:281-7.  Back to cited text no. 102
Dey SS, Singh AK, Chandel D, Behera TK. Genetic diversity of bitter gourd (Momordica charantia L.) genotypes revealed by RAPD markers and agronomic traits. Sci Hortic 2006;109:21-8.  Back to cited text no. 103
Luan S, Chiang TY, Gong X. High genetic diversity vs. low genetic differentiation in Nouelia insignis (Asteraceae), a narrowly distributed and endemic species in China, revealed by ISSR fingerprinting. Ann Bot 2006;98:583-9.  Back to cited text no. 104
Hess J, Kadereit JW, Vargas P. The colonization history of Olea europaea L. in Macaronesia based on internal transcribed spacer 1 (ITS-1) sequences, randomly amplified polymorphic DNAs (RAPD), and intersimple sequence repeats (ISSR). Mol Ecol 2000;9:857-68.  Back to cited text no. 105
Carolan JC, Hook IL, Walsh JJ, Hodkinson TR. Using AFLP markers for species differentiation and assessment of genetic variability of in vitro-cultured Papaver Bracteatum (Section Oxytona). In Vitro Cell Dev Biol Plant 2002;38:300-7.  Back to cited text no. 106
Chaurasia AK, Subramaniam VR, Krishna B, Sane PV. RAPD based genetic variability among cultivated varieties of Aonla (Indian Gooseberry, Phyllanthus emblica L.) Physiol Mol Biol Plants 2009;15:169-73.  Back to cited text no. 107
Barazani O, Dudai N, Golan-Goldhirsh A. Comparison of mediterranean Pistacia lentiscus genotypes by random amplified polymorphic DNA, chemical and morphological analyses. J Chem Ecol 2003;29:1939-52.  Back to cited text no. 108
Abhyankar G, Reddy VD, Giri CC, Rao KV, Lakshmi VV, Prabhakar S, et al. Amplified fragment length polymorphism and metabolomic profiles of hairy roots of Psoralea corylifolia L. Phytochemistry 2005;66:2441-57.  Back to cited text no. 109
Hu Y, Wang L, Xie X, Yang J, Li Y, Zhang H. Genetic diversity of wild populations of Rheum tanguticum endemic to China as revealed by ISSR analysis. Biochem Syst Ecol 2010;38:264-74.  Back to cited text no. 110
Wang Y, Li XE, Li XD, Qi JJ, Sun P, Zhou LL. Analysis of genetic diversity of wild Rehmannia glutinosa by using RAPD and ISSR markers. Zhongguo Zhong Yao Za Zhi 2008;33:2591-5.  Back to cited text no. 111
Hu Y, Wang L, Xie X, Yang J, Li Y, Zhang H. Genetic diversity of wild populations of Rheum tanguticum endemic to China as revealed by ISSR analysis. Biochem Syst Ecol 2010;38:264-74.  Back to cited text no. 112
Xia T, Chen S, Chen S, Zhang D, Gao Q, Ge X. ISSR analysis of genetic diversity of the Qinghai-Tibet plateau endemic Rhodiola chrysanthemifolia (Crassulaceae). Biochem Syst Ecol 2007;35:209-14.  Back to cited text no. 113
Elameen A, Klemsdal SS, Dragland SM, Fjellheim S, Rognli OA. Genetic diversity in a germplasm collection of roseroot (Rhodiola rosea) in Norway studied by AFLP. Biochem Syst Ecol 2008;36:706-15.  Back to cited text no. 114
Zhao F, Liu F, Liu J, Put O, Ang J, Duan D. Genetic structure analysis of natural Sargassum muticum (Fucales, Phaeophyta) populations using RAPD and ISSR markers. Appl Phycol 2008;20:191-8.  Back to cited text no. 115
Kochieva EZ, Khussein IZ, Legkobit MP, Khadeeva NV. The detection of genome polymorphism in Stachys species using RAPD. Russ J Genet 2002;38:516-20.  Back to cited text no. 116
Muchugi A, Muluvi GM, Kindt R, Kadu CA, Simons AJ, Jamnadass RH. Genetic structuring of important medicinal species of genus Warburgia as revealed by AFLP analysis. Tree Genet Genome 2008;4:787-95.  Back to cited text no. 117
Dhar RS, Verma V, Suri KA, Sangwan RS, Satti NK, Kumar A, et al. Phytochemical and genetic analysis in selected chemotypes of Withania somnifera. Phytochemistry 2006;67:2269-76.  Back to cited text no. 118


  [Figure 1], [Figure 2]

  [Table 1], [Table 2]

This article has been cited by
1 Molecular authentication of Trichosanthes species traded as “Patola:” An ayurvedic drug resource
Bhagyashri Kumbhalkar,Anuradha Upadhye,Shubhada Tamhankar
Pharmacognosy Magazine. 2018; 14(55): 52
[Pubmed] | [DOI]


Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

  In this article
Molecular Marker
Basic Molecular ...
Amplified Fragme...
Nonpolymerase Ch...
Microsatellites ...
Advances in Mole...
RNA-based Molecu...
DNA Barcoding
Article Figures
Article Tables

 Article Access Statistics
    PDF Downloaded423    
    Comments [Add]    
    Cited by others 1    

Recommend this journal