Effects of lemon balm (Melissa officinalis L.) extract on germination and seedling growth of six plants

The n-hexane-, acetone- and water-soluble fractions obtained from an aqueous acetone extract of lemon balm (Melissa officinalis L.) shoots inhibited the germination and the growth of roots and shoots of cockscomb (Amaranthus caudatus L.), cress (Lepidium sativum L.), crabgrass (Digitaria sanguinalis L.), timothy (Phleum pratense L.), lettuce (Lactuca sativa L.) and ryegrass (Lolium multiflorum Lam.). The inhibitory activity of the water-soluble fraction was the greatest, followed by that of acetone- and n-hexane-soluble fractions in all bioassays. The effectiveness of these fractions on the roots was greater than that of the shoots of the test plants. Significant reductions in the germination and growth of the roots and shoots were observed as the extract concentration increased. Such rate-dependent responses of the test plants to the fractions suggest that each fraction might contain allelochemical(s), but that the greatest potential was in the water-soluble fraction.     

Enzymes of phenylpropanoid metabolism in the important medicinal plant Melissa officinalis L.

Lemon balm (Melissa officinalis, Lamiaceae) is a well-known medicinal plant. Amongst the biologically active ingredients are a number of phenolic compounds, the most prominent of which is rosmarinic acid. To obtain better knowledge of the biosynthesis of these phenolic compounds, two enzymes of the general phenylpropanoid pathway, phenylalanine ammonia-lyase (PAL) and 4-coumarate:coenzyme A-ligase (4CL), were investigated in suspension cultures of lemon balm. MoPAL1 and Mo4CL1 cDNAs were cloned and heterologously expressed in Escherichia coli and the enzymes characterised. Expression analysis of both genes showed a correlation with the enzyme activities and rosmarinic acid content during a cultivation period of the suspension culture. Southern-blot analysis suggested the presence of most probably two gene copies in the M. officinalis genome of both PAL and 4CL. The genomic DNA sequences of MoPAL1 and Mo4CL1 were amplified and sequenced. MoPAL1 contains one phase 2 intron of 836 bp at a conserved site, whilst Mo4CL1 was devoid of introns.     

The effect of triacontanol on micropropagation of balm, Melissa officinalis L.

 Triacontanol, a long-chain primary alcohol was found to be an effective growth regulator in the micropropagation of balm, Melissa officinalis. In both the multiplication and the rooting phase, concentrations of 2, 5, 10 and 20 μg triacontanol per liter were applied. After 4 weeks of culture, the fresh weight of shoots was measured in the multiplication phase and root formation, photosynthetic activity, chlorophyll content and the fresh and dry weights of shoots were analyzed in the root induction phase. In the multiplication phase, 5 μg/l triacontanol was found to be the optimal concentration, while in the rooting phase 2 μg/l was the most effective. Triacontanol increased the number and length of roots, and it enhanced shoot growth, fresh weight, and the chlorophyll content, but it had no effect on the dry weight and the photosynthetic activity of the plants. Results of our work demonstrate that triacontanol can be applied as an effective growth regulator in the tissue culture of balm. Received: 3 December 1997 / Revised: 24 February 1998 / Accepted: 26 February 1999     

Development and characterisation of microsatellite markers for the medicinal plant Scutellaria baicalensis (Lamiaceae)

Scutellaria baicalensis is a popular medicinal plant that is on the verge of extinction due to uncontrolled harvesting, habitat destruction and deterioration of its ecosystem. We isolated and characterised 21 microsatellite loci in this species. Ninety-four individuals from six populations were used to test the polymorphism of the microsatellite loci. The number of alleles per locus ranged from 1 to 13, with a mean of 7.2. Observed and expected heterozygosities varied from 0.000 to 1.000 and 0.000 to 0.938, respectively. Among these new microsatellite markers, only two loci showed significant deviation from Hardy–Weinberg equilibrium. No locus pairs showed significant linkage disequilibrium. The 21 primer pairs were tested in other Scutellaria species. Most of these primer pairs worked successfully, except for Scut18. These new microsatellite markers could be applied to investigate the genetic diversity and population genetic structure of S. baicalensis and its closely related species.     

DNA isolation protocol for red seaweed (rhodophyta)

We report a DNA isolation protocol for red seaweed. The method is a modification of the Dellaporta et al. (1983) protocol for land plants. Our simplified version can be used to process large sample numbers and to minimise polysaccharide co-isolation. The protocol was applied to 12 red seaweed species as well as one green alga and one land plant. The protocol yields about 5 μg of high molecular weight DNA from 10 mg of dried material, with no RNA. No sign of degradation was observed after agarose gel electrophoresis for both freshly extracted DNA and DNA stored for 18 months at 4°C. DNA isolated by our protocol was suitable for genomic library construction (tested for one species), endonuclease restriction, and PCR amplification for all species.     

Simple and rapid protocol for the isolation of PCR-amplifiable DNA from medicinal plants

Medicinal plant species has a valuable economic importance because of its usage as pharmaceuticals, nutritional, as well as its use in popular medication. For DNA-based techniques, nanogram quantities of the purified DNA are requisite to amplify and yield sufficient amounts of PCR products. SDS-based DNA isolation method was used to extract DNA from 11 species of different aromatic and medicinal plants collected from Saudi Arabia. Three hundred milligrams of fresh shredded plant material was necessary. The DNA purity was further confirmed by agarose gel, restriction endonuclease digestion and microsatellite primed-polymerase chain reaction (MP-PCR). DNA yields ranged from 10-20 μg (in 100-μL elution volumes) from all plant material evaluated. The DNA obtained was free of any contaminating proteins, polysaccharides and colored pigments. The extracted genomic DNA was found suitable for restriction digestion and PCR amplification. Our experimental procedure provides an easy, suitable, non-toxic, cheap, and quick process for the amplification of DNA from medical plant tissue.     

Factors influencing seed germination of medicinal plant Salvia aegyptiaca L. (Lamiaceae)

Salvia aegyptiaca is a xerophytic perennial herb belongs to the Lamiaceae family commonly used for medicinal purposes. Laboratory experiments were carried out to assess the effects of temperature and salinity on seed germination and recovery responses after transferring to distilled water. Temperatures between 10 and 40 °C seem to be favourable for the germination of this species. Germination was inhibited by either an increase or decrease in temperature from the optimum (30 °C). The highest germination percentages were obtained at 0 mM NaCl; however, the increase of solution osmolalities progressively inhibited seed germination. The germination rate decreased with an increase in salinity for most of tested temperatures, but comparatively higher rates were obtained at 30 °C. Salt stress decreased both the percentage and the rate of germination. An interaction between salinity and temperature yielded no germination at 300 mM NaCl. By experimental transfer to distilled water, S. aegyptiaca seeds that were exposed to moderately saline conditions recovered and keep their ability to germinate mostly at low temperatures. At 300 mM NaCl, germination recovery decreased with increasing temperature and it was completely inhibited at 40 °C.     

DNA isolation protocol for seaweeds

We report a DNA isolation protocol for red seaweeds. Recovering DNA of high quality and quantity is a prerequisite for ensuring suitable applications, such as polymerase chain reaction (PCR), restriction fragment length polymorphism (RFLP) analysis, and sequencing. Isolation of DNA from seaweeds has proven difficult because of coprecipitation of polysaccharides. Our process minimizes this contamination, which is mostly due to the highly hydrocolloidal content of algal cell walls. This protocol, using 2 steps, is based on a preliminary enzymatic digestion of cell wall with specific enzymes (Novozymes) followed by centrifugation, allowing isolation of DNA on the pellet. This provides a higher yield of DNA, in the range of 40 μg (Palmaria palmata) and 18 μg (Gracilaria verrucosa) from 50 mg of fresh frozen pellet.     

A modified protocol for rapid DNA isolation from plant tissues using cetyltrimethylammonium bromide

We describe a modification of the DNA extraction method, in which cetyltrimethylammonium bromide (CTAB) is used to extract nucleic acids from plant tissues. In contrast to the original method, the modified CTAB procedure is faster, omits the selective precipitation and CsCl gradient steps, uses less expensive and toxic reagents, requires only inexpensive laboratory equipment and is more readily adapted to high-throughput DNA extraction. This protocol yields approximately 5-30 microg of total DNA from 200 mg of tissue fresh weight, depending on plant species and tissue source. It can be completed in as little as 5-6 h.     

Diversity,phytochemical and medicinal potential of the genus Ocimum L. (Lamiaceae)

Phytochemistry Reviews - Ocimum, commonly known as Tulsi, is a huge genus within family Lamiaceae, comprising about 64 species of annual to perennial aromatic medicinal herbs with a long history of...     

DNA isolation methods for medicinal and aromatic plants

Several protocols described for plant DNA isolation fail to produce good quality DNA from medicinal herbs and aromatic plants. These plants contain exceptionally high amounts of secondary metabolites that interfere with DNA isolation. To address this problem, we developed 2 DNA isolation methods for sundew and tarragon that produce DNA suitable for molecular biological applications. One of the methods also is applicable for milfoil and Siberian ginseng.     

Magnetic, microplate-format plasmid isolation protocol for high-yield, sequencing-grade DNA

We have developed a rapid, microplate-format plasmid isolation procedure to purify sequencing-grade DNA templates for high-throughput DNA sequencing operations. A modified lysozyme/boiling method is used to produce a plasmid-containing supernatant that is then purified by iron bead capture. After binding, the beads are pelleted in a magnetic field, washed and the DNA eluted in water. The method yields up to 10 micrograms plasmid DNA from a 1-mL overnight culture in a deep-well microplate. The procedure is suitable for large-scale experiments, amenable to automation and does not require expensive reagents or equipment. The entire protocol can be completed in as little as 2 h, and one technician with a 96-well pipetting station can process up to 48 plates per day. This protocol is ideal for any high-throughput operation in which template quantity, quality and reproducibility are of primary importance.     

A simple protocol for isolation of fungal DNA

Genomic DNA was isolated from as little as 2 mg dry biomass of Magnaporthe grisea by microwave treatment within 30 s. The quantity of DNA was good enough for PCR analysis and Dot blot hybridization. This technique can be used for various studies, such as DNA fingerprinting to study the population structure of the phytopathogen in different regions, and for a quick screening of M. grisea transformants.     

Volatile components of whole and different plant parts of bastard balm (Melittis melissophyllum L., Lamiaceae) collected in Central Italy and Slovakia

The aim of this work was to trap the volatiles released from whole frozen and dry aerial parts, and, separately, from different organs (leaves, stems, corolla and calyx) of bastard balm (Melittis melissophyllum L., Lamiaceae) populations collected in Italy and Slovakia by HS-SPME, and to identify the headspace constituents responsible for the characteristic aroma impression by GC/FID and GC/MS techniques. Among more than 100 volatile components detected, the C(8) alcohol oct-1-en-3-ol, responsible for the typical mushroom-like odor, and the phenolic coumarin, with a characteristic sweet and creamy vanilla bean odor, played a major role in the aroma of whole aerial parts and different plant organ samples. In particular, dry calyx parts could be proposed as flavoring agent in food products as mushroom aroma enhancer. Multivariate chemometric techniques, such as cluster analysis and principal component analysis, were used to characterize the sample populations according to the geographical origin and processing of plant material.     

Micropropagation and cryopreservation by vitrification of the Spanish endemic medicinal plant Sideritis leucantha Cav. subsp. leucantha (Lamiaceae)

In Vitro Cellular & Developmental Biology - Plant - Sideritis leucantha Cav. subsp. leucantha is a Spanish endemic medicinal plant that faces conservation problems. Therefore, this work is...     

DNA barcoding of medicinal plant material for identification

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