Development of an allele-specific PCR (AS-PCR) method for identifying high-methyl eugenol-containing purple Tulsi (Ocimum tenuiflorum L.) in market samples.

BACKGROUND: Ocimum tenuiflorum L. is a highly traded medicinal with several therapeutic values. Green Tulsi and purple Tulsi are two subtypes in O. tenuiflorum and both have the same medicinal properties. Recent reports have revealed that purple Tulsi contains higher quantities of methyl eugenol (ME), which is moderately toxic and potentially carcinogenic. Therefore, we developed an allele-specific PCR (AS-PCR) method to distinguish the green and purple Tulsi. METHODS AND RESULT: Using the green Tulsi as a reference, 12 single nucleotide polymorphisms (SNPs) and 10 insertions/deletions (InDels) were identified in the chloroplast genome of the purple Tulsi. The C > T SNP at the 1,26,029 position in the ycf1 gene was selected for the development of the AS-PCR method. The primers were designed to amplify 521 bp and 291 bp fragments specific to green and purple Tulsi, respectively. This AS-PCR method was validated in 10 accessions from each subtype and subsequently verified using Sanger sequencing. Subsequently, 30 Tulsi powder samples collected from the market were subjected to molecular identification by AS-PCR. The results showed that 80% of the samples were purple Tulsi, and only 3.5% were green Tulsi. About 10% of the samples were a mixture of both green and purple Tulsi. Two samples (6.5%) did not contain O. tenuiflorum and were identified as O. gratissimum. CONCLUSION: The market samples of Tulsi were predominantly derived from purple Tulsi. The AS-PCR method will be helpful for quality control and market surveillance of Tulsi herbal powders.

Multi-locus DNA barcoding identifies matK as a suitable marker for species identification in Hibiscus L.

The genus Hibiscus L. includes several taxa of medicinal value and species used for the extraction of natural dyes. These applications require the use of authentic plant materials. DNA barcoding is a molecular method for species identification, which helps in reliable authentication by using one or more DNA barcode marker. In this study, we have collected 44 accessions, representing 16 species of Hibiscus, distributed in the southern peninsular India, to evaluate the discriminatory power of the two core barcodes rbcLa and matK together with the suggested additional regions trnH-psbA and ITS2. No intraspecies divergence was observed among the accessions studied. Interspecies divergence was 0%-9.6% with individual markers, which increased to 0%-12.5% and 0.8%-20.3% when using two- and three-marker combinations, respectively. Differentiation of all the species of Hibiscus was possible with the matK DNA barcode marker. Also, in two-marker combinations, only those combinations with matK differentiated all the species. Though all the three-marker combinations showed 100% species differentiation, species resolution was consistently better when the matK marker formed part of the combination. These results clearly showed that matK is more suitable when compared to rbcLa, trnH-psbA, and ITS2 for species identification in Hibiscus.

Chloroplast genome of Ocimum basilicum var. purpurascens Bentham 1830 (Lamiaceae).

Ocimum basilicum var. purpurascens Bentham 1830 (Red Rubin Basil) is an aromatic herb belonging to the family Lamiaceae and is known for its medicinal uses. It is commonly used in traditional medicine to treat cardiovascular diseases and obesity. It possesses anti-inflammatory, antioxidant, antifungal, and anti-spasmodic properties. In our recent study, we assembled the chloroplast genome sequence of O. basilicum var. purpurascens using Illumina paired-end sequencing technology. The assembled chloroplast genome was 152,407 base pairs (bp), inclusive of a large single-copy (LSC) region accounting for 83,409 bp and a small single-copy (SSC) region spanning 17,604 bp. Two inverted repeats (IRs) interspersed these regions, each 25,697 bp long. The chloroplast genome harbored 132 genes, comprising 88 protein-coding genes, 36 transfer RNA (tRNA), and eight rRNA genes. Among these, nine genes encompassed a single intron, two presented with two introns, with the remaining devoid of any introns. The overall GC content of the chloroplast genome was determined to be 38%. The GC content in the LSC, SSC, and IR regions was 35.9%, 31.6%, and 43.1%, respectively. Our phylogenetic exploration of the chloroplast genomes elucidated that O. basilicum var. purpurascens exhibits close genetic affinity with O. basilicum var. basilicum and other constituents of the Ocimum genus within the Lamiaceae family.

The complete chloroplast genome of Ocimum americanum Linnaeus 1755 and phylogenetic analysis among the Lamiaceae family.

Ocimum americanum Linnaeus 1755 (Lemon basil) is an essential medicinal species in the Ocimum genus. Its leaf decoction is traditionally used to treat diabetes, constipation, diarrhea, piles, and dysentery. The essential oils from this species have intense fungicidal activity. The complete chloroplast genome sequence of O. americanum was assembled from Illumina paired-end sequencing data. The O. americanum chloroplast genome was 152,460 bp in length, containing a large single copy (LSC) region of 83,459 bp and a small single copy (SSC) region of 17,607 bp, separated by a pair of inverted repeats (IRs) of 25,697 bp. The genome contained 134 unique genes, including 89 protein-coding, 37 tRNA, and eight rRNA genes. Among them, nine genes had a single intron, and two genes contained two introns. The overall GC content of the chloroplast genome was 38%, while the corresponding values of LSC, SSC, and IR regions were 35.8%, 31.7%, and 43.1%, respectively. In the phylogenetic analysis, all the Ocimum species formed a group closely related to Plectranthus barbatus. O. americanum was more closely related to O. gratissimum and O. basilicum than the other species of Ocimum included in this study.

The complete chloroplast genome of Phyla nodiflora (Linnaeus) Greene (1899) from the Verbenaceae family and its phylogenetic analysis.

Phyla nodiflora (Linnaeus) Greene (1899) is a perennial creeping herb belonging to the family Verbenaceae. It has numerous pharmacological properties, including anti-dandruff, anti-inflammatory, anti-oxidant, anti-melanogenesis, anti-hypertensive, and anti-hyperuricemic properties. We generated the complete chloroplast genome sequence of P. nodiflora using Illumina paired-end sequencing data. The P. nodiflora chloroplast genome is 154,341 bp in length, containing a large single copy (LSC) region of 85,185 bp and a small single copy (SSC) region of 17,222 bp, separated by a pair of inverted repeats (IRs) of 25,967 bp. The genome contained 128 genes, including 86 protein-coding, 34 tRNA, and eight rRNA genes. Six genes had one intron, one gene had two introns, and the others did not have an intron. Overall GC content of the chloroplast genome was 39%, while those of LSC, SSC, and IR regions were 38.2%, 33.7%, and 44%, respectively. Phylogenetic analysis of the chloroplast genome revealed that P. nodiflora is closely related to the other species from Verbenaceae.

The complete chloroplast genome of Ocimum basilicum L. var. basilicum (Lamiaceae) and its phylogenetic analysis.

Ocimum basilicum L. var. basilicum (Sweet Basil) is an aromatic herb belonging to the family Lamiaceae and is known for its medicinal uses. It is commonly used in traditional medicine for its therapeutic value, including anti-allergic, anti-inflammatory, antioxidant, antitumor, and antimicrobial properties. In this study, we generated the complete chloroplast genome sequence of O. basilicum var. basilicum using Illumina paired-end sequencing data. The chloroplast genome was 152,407 bp in length, containing a large single-copy (LSC) region of 83,409 bp and a small single-copy region (SSC) of 17,604 bp, separated by a pair of inverted repeats (IRs) of 25,697 bp. The genome contained 134 genes, including 89 protein-coding, 37 tRNA, and eight rRNA genes. Nine genes had one intron, two genes had two introns, and others did not have any intron. Overall GC content of the chloroplast genome was 38%, while that of LSC, SSC, and IR regions was 35.9%, 31.6%, and 43.1%, respectively. Phylogenetic analysis of the chloroplast genomes revealed that O. basilicum var. basilicum was closely related to Ocimum basilicum from the Ocimum species.

The complete chloroplast genome of Ocimum tenuiflorum L. subtype Rama Tulsi and its phylogenetic analysis.

Ocimum tenuiflorum L. subtype Rama Tulsi is an important aromatic perennial herb. It belongs to the family of Lamiaceae. In this study, the complete chloroplast genome sequence of O. tenuiflorum subtype Rama Tulsi was assembled and annotated using Illumina paired-end sequencing data. The length of the complete circular chloroplast genome was 151,722 bp. It comprises an inverted repeat (IR) region with a repeat length of 25,677 bp, a large single-copy (LSC) region of 82,781 bp, and a small single-copy (SSC) region of 17,587 bp. The GC content of complete chloroplast genome, LSC, SSC, IR regions is 37.9%, 36.0%, 31.8%, and 43.1%, respectively. The chloroplast genome contains 134 genes, including 88 protein-coding genes, 38 transfer RNA genes, and eight ribosomal RNA genes. Phylogenetic analysis with the complete chloroplast genomes of other related species revealed that the O. tenuiflorum L. subtype Rama Tulsi is fully resolved in a clade with other Ocimum species classified under the Lamiaceae family.

The complete chloroplast genome and phylogenetic analysis of Ocimum kilimandscharicum Gurke (Camphor Basil) from India.

Ocimum kilimandscharicum Gurke commonly known as Camphor Basil, is a medicinal plant species that belongs to the Lamiaceae family. Here, the sequencing and characterization of complete chloroplast genome sequence of O. kilimandscharicum is reported for the first time using Illumina paired-end sequencing data. The size of the chloroplast (cp) genome is 151,741 bp in length, with a large single-copy (LSC) region of 82,882 bp and a small single-copy (SSC) region of 17,587 bp, separated by a pair of 25,636 bp inverted repeat (IR) regions. There are 135 predicted genes, including 90 protein-coding genes, 37 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes in the genome, and the overall GC content of the genome is 37.9%. The phylogenetic analysis based on the chloroplast genome data indicated that O. kilimandscharicum is closer to O. tenuiflorum and clustered to other Ocimum species in Lamiaceae.

The complete chloroplast genome of Ocimum gratissimum from India - a medicinal plant in the Lamiaceae.

Ocimum gratissimum L. is an important medicinal species with several therapeutic applications. It is used in traditional medicine as a single drug and in formulations. We generated the complete chloroplast genome sequence of O. gratissimum by using Illumina paired-end sequencing data. The O. gratissimum chloroplast genome is 152,469 bp in length, containing a large single copy (LSC) region of 83,614 bp and a small single copy region (SSC) of 17,607 bp, separated by a pair of inverted repeats (IRs) of 25,624 bp. The genome contains 138 unique genes, including 85 protein-coding, 45 tRNA, and eight rRNA genes. Among them, six genes have one intron each, and two genes contain two introns. The overall GC content of the chloroplast genome is 37.8%, while the corresponding values of LSC, SSC, and IR regions are 35.6%, 31.7%, and 43.2%, respectively. Phylogenetic analysis with the complete chloroplast genomes of other related species revealed that O. gratissimum is fully resolved in a clade with other Ocimum species classified to the family Lamiaceae.

The complete chloroplast genome of Ocimum tenuiflorum L. subtype Krishna Tulsi and its phylogenetic analysis.

The complete chloroplast genome (cp) of Ocimum tenuiflorum L. subtype Krishna Tulsi was sequenced and assembled using Illumina paired-end sequencing data. The cp genome is 151,758 bp in length, including a large single copy (LSC) region of 82,794 bp, a small single-copy region (SSC) of 17,592 bp, and a pair of inverted repeated (IR) region of 25,686 bp. The cp genome of Krishna Tulsi encodes 129 genes, including 90 protein-coding, 31 transfer RNA (tRNA), and eight ribosomal RNA (rRNA) genes. While the overall GC content was 37.9%, it is 36.0%, 31.8%, and 43.1% in the LSC, SSC, and IR regions, respectively. Phylogenetic analysis based on chloroplast genome sequences of 17 species from Lamiaceae revealed that O. tenuiflorum subtype Krishna Tulsi is clustered with other Ocimum species, and forms a clade with genera from family Lamiaceae.

A reference-grade genome identifies salt-tolerance genes from the salt-secreting mangrove species Avicennia marina.

Water scarcity and salinity are major challenges facing agriculture today, which can be addressed by engineering plants to grow in the boundless seawater. Understanding the mangrove plants at the molecular level will be necessary for developing such highly salt-tolerant agricultural crops. With this objective, we sequenced the genome of a salt-secreting and extraordinarily salt-tolerant mangrove species, Avicennia marina, that grows optimally in 75% seawater and tolerates >250% seawater. Our reference-grade ~457 Mb genome contains 31 scaffolds corresponding to its chromosomes. We identified 31,477 protein-coding genes and a salinome consisting of 3246 salinity-responsive genes and homologs of 614 experimentally validated salinity tolerance genes. The salinome provides a strong foundation to understand the molecular mechanisms of salinity tolerance in plants and breeding crops suitable for seawater farming.