DNA barcoding technique is a useful tool for the identification of plant and animal species using a short and standard sequence of the genome. In the present study, this method was used to identify four plant species including Calendula persica C. A. Mey., Silybum marianum (L. ) Gaertn., Satureja mutica Fisch. & C. A. Mey., and Malva neglecta Wallr. from the eastern Golestan province. The DNA was extracted by CTAB method and the PCR was performed with the primers designed based on the rbcL and trnH-psbA chloroplast barcodes and ITS nuclear barcode. The results of sequences were matched with the information in the NCBI database. The results showed that the all three barcodes were suitable for the samples studied due to their high resolution, low SNP number, and comprehensiveness in most species. Also, the barcodes comparison of the species collected from the rangelands and perfumeries showd that some plant species that are offered in the perfumeries are different from the plants that the natives use as medicine. It could be mentioned that the mistakes possibility in the medicinal plants offered in the perfumeries is undeniable. Therefore, the study on the other plant species in the perfumeries by the DNA barcoding method could be recommended as a necessity.

Background and Objectives: Jujube (Ziziphus jujuba Mill. ) is one of the most important medicinal plants belong to the Rhamnaceae family. It is important in the pharmaceutical industry. Genetic diversity of different species of Z. jujuba in Iran in 2007 and 2006 classified the sex of 19 species by simultaneous analysis of morphological traits and molecular methods. The main purpose of this study is DNA barcoding of different ecotypes of Z. jujuba in South Khorasan, Iran using two chloroplast genes (rbcL and matK). Materials and Methods: The 25 numbers of the ecotypes of this species from 6 different provinces of Iran were planted in South Khorasan, were assessed. Two close relatives of the same family (Sangoisorba sp., Rosa sp. ) were also used as external groups. Purification (protein and polysaccharide refining) was performed using the manual method. The gel was then stained with ethidium bromide and the DNA quality was estimated using the agarose gel electrophoresis results. The chloroplast genes from the DNA extracted were amplified using the PCR technique. All sequences obtained from forward and reverse reading in this study to produce the final sequence using appropriate software were assembled. Results: The DNA barcode of each species was performed for fast, accurate and automatic identification of the species and all the sequences obtained from this study were sent to the NCBI (National Center for Biotechnology Information) and submitted. The results show that there is the greatest genetic distance between jujube ecotypes and two samples from the outgroup and no significant genetic diversity between different jujube ecotypes and morphological differences are due to ecological conditions. However, the analysis of gene combination data provided more informational insights than the separate analysis of sequence data. According to the results, the highest diversity was observed in the ecotypes of South Khorasan. Conclusion: The results of pairwise distance and haplotype networks showed the most variety among different ecotypes belonged to the different areas of South Khorasan province. Also, Soth Khorasan can be considered as the origin of this species in Iran. Previous studies have fully confirmed the results of this study. However, given the economic importance of jujube plants in the world, Iran and especially South Khorasan, it is recommended to use more samples and more genes, especially nuclear genes, to study more closely the genetic similarities and differences of these ecotypes.

The plant family Lamiaceae contains several species with potential therapeutic activity due to their essential oils. In this study, the chloroplast genes of two species of the Lamiaceae family, Thymus syriacus and Salvia nemorosa, were sequenced to make the identification of these two species quickly and accurately. The genes matK and rbcL were used for sequencing Thymus syriacus and matK, trnL-F for Salvia nemorosa. At first, genomic DNA was extracted from the leaves of these two species, then the sequences of the desired chloroplast genes were amplified and eventually sequenced. The sequence of the obtained chloroplast genes can be used in other phylogenetic and evolutionary studies to compare the species and could therefore provide more accurate results within a shorter time especially in large-scale studies. The results shows that chloroplast regions as matK, rbcL and trnL-F can be used as DNA barcodes for identification of Salvia nemorosa and Thymus syriacus species.

Background and objectives: As a member of the Lamiaceae family native to Europe, Asia, and North Africa, Vitex agnus-castus is a popular medicinal plant. Despite much research, it is always essential to verify the safety of this valuable species, as it is among the world's best-selling plants. Several species in the genus Vitex do not exhibit recognizable morphologies, confusing their identification. The medicinal use of V. agnus-castus differs from that of other species. For effective pharmaceutical performance, the identification of this species is essential. This study used morphological, micromorphological, and molecular approaches to identify this medicinal plant.Methodology: A total of 17 populations of the target species were investigated in Maraveh Tappeh, a city in the Golestan province located in the eastern region. Pollen samples were collected directly from the natural habitat of the target area. After acetolysis, 30-40 pollen grains were photographed from the polar and equatorial views with a light microscope and 40 and 100 magnification and with the help of a Canon digital camera. In order to study the seed morphology, fully ripe fruits were collected from each studied population during the fruiting season. Twenty seeds from each population were kept for photography with light microscopy and Scanning electron microscopy (SEM). Electron micrographs were prepared from suitable seeds and pollen at the Razi Metallurgical Research Center (RMRC) using an SEM electron microscope. Leaf cells from herbarium samples were extracted from DNA using a DNA extraction kit. Plastid trnL-trnF sequences and nrDNA ITS region sequences were used as barcodes. Polymerase chain reaction (PCR) was performed in 20 microliters with desired primers and a specific temperature program in a thermocycler. After performing the polymerase chain reaction, in order to ensure the amplification of fragments, the final product was electrophoresed. Strong single bands were sent to Codon Genetics Company in Tehran for sequencing.Results: Pollen grains of all species are small (12-28 micrometers). According to Ertman's classification, Ghazan ghayeh pollen grains are prolate spheroidal (elongated spherical), and Ghoshe Tappeh pollen grains are subprolate (semi-elongated). All pollen grains are tricolporate. Ghazan ghayeh ornamentation is micro-perforated, while Ghoshe Tappeh ornamentations are reticulated-micro-perforated. With an average length of 3.84 mm and width of 1.52 mm, Ghazan ghayeh seeds are the largest. They are all almond-shaped, but in the Ghazan Ghayeh population, the outer surface of the seed is hollow; in the Ghoshe Tappeh population, the surface is wrinkled and striated. The trnL-trnF gene locus was found to have a multiplication success rate of 85% in the examined plants. There were 516 nucleotides in the amplified fragment in this species. For final registration, the sequence was sent to the GenBank. A 675 nucleotide fragment amplified from this species' ITS marker was sequenced, and the chromatograms were compared with the NCBI database. The gene bank has been notified of the sequence determination. The results showed the highest similarity (98%) with the Vitex agnus-castus species reported from America.Conclusion: Local herbal medicines are gaining popularity in different countries and play a significant role in treating diseases today. In order to make medicinal plants more accessible, we need to pay more attention to trust, marketing, and consumption. Barcoding and molecular approaches serve this purpose effectively. To compare plant samples available on the market with natural medicinal plants using DNA barcoding of correctly identified medicinal species. Hence, the sequences used in this study are essential for barcoding Vitex agnus-castus. The medicinal species has been correctly identified and can be used as a standard for evaluating the plants available on the market.

DNA barcoding is a simple way to identify species using a very short genetic sequence from a standard part of the genome. This technique used to identify eight medicinal plants collected from the Ardabil province. DNA extraction was performed by modified CTAB method and PCR was performed with primers designed based on rbcL, trnH-psbA, matKChloroplast barcodes and ITS nuclear barcode. Then, PCR products purified and sequenced. The percentage of amplification and sequencing success were assumed in samples respectively, 87 and 62, 75 and 37, 62 and 12, 75 and 37. The sequences were blasted with samples existed in NCBI database and Bioinformatics analyses were performed. In phylogenetic tree, the species belonging to the same genus were separated from other genus based on rbcL and trnH-psbA barcode sequences. Also, in ITS barcode only G. glabra organized with plants from same genus. In this study, barcoding of L. ledebourii with rbcL was done for the first time. SNPs were counted for barcodes of rbcL (less than 30), trnH-psbA(less than100), ITS (more than 200) and matK (less than 20). Thus, rbcL barcode due to high separation ability, low number of SNPs and universality in most species, was introduced as the best barcode. However, trnH-psbA and ITS barcodes due to related problem with direct sequencing of PCR products and lack of access to high quality sequences were identified as complementary barcodes. MatK barcode is not recommended for these samples because of the low ability of amplification and sequencing.

This study included 81 samples which belonged to 13 speices from Myriophyllum genus. Both nrDNA ITS1 and ITS2 and cpDNA matK, rbcL and trnH-psbA loci were used for diagnose of invasive from native plant. The nrDNA ITS1and ITS2 data proved highly variable and could differentiate between all but had low PCR amplification success, based on these result they would not recommend as a suitable barcode in Myriophyllum genus. Although matK had high amplification success but had low sequencing success which could not be ideal barcode among studied species. Based on result Non-coding trnH-psbA spacer region and a portion of the coding rbcL gene are recommended as ideal barcode that provide the necessary universality and species discrimination among Myriophyllum species. a limited investment in DNA barcoding can generate an identification tool for plant material, specifically useful for cases where morphology is inadequate to assess species identity.

The objective of this study was to detect the level of SNP variations of rbcL gene sequences among and withinPrunus species including 17 locally cultivated and wild relatives of Prunus, and two species of the subfamily Maloideae (Malus domestica and Pyrus communis), as out groups. The rbc L sequences were amplified, sequenced, and aligned to determine Single Nucleotide Polymorphisms (SNPs). TherbcL gene tree of the samples showed two main clusters. The first included the outgroup taxa (M. domestica and P. communis); and all Prunus samples in the second cluster including Prunus armeniaca, which separated in a subcluster. Our results indicate that rbcL gene sequence analysis provides a well-defined tool to study relationships within and amongPrunus species, and can be successfully used in constructing reliable phylogenetic tree for Prunus accessions.

Background: Genetic manipulation of chloroplast in higher plants offers a number of unique prerogatives, including; undesirable of pleiotropic genome and gene silencing effects and also use as an important agronomic trait for producing essential biomaterials and industrial enzymes. In order to manipulate chloroplast genome, specific vectors are required. These vectors can be transformed and expressed in Escherichia coli due to the same evolutionary origin of bacteria and chloroplasts.Objectives: The aim of the present study was to construct chloroplast vector specified for spinach and assessing the chloroplast regulatory elements in a prokaryotic expression host, E. coli.Materials and Methods: Flanking sequences (INSL+INSR) were isolated by PCR from the spinach chloroplast genome and blunt-end ligated into the PvuII site of pUC19 vector to form an intermediate vector, pUCINS. Then the selectable marker cassette (including aadA gene, Prrn promoter and rbcL terminator) was isolated via PCR and blunt-end cloned into the unique PvuII site of pUCINS to make the final chloroplast vector, named pCSI.Results: The constructed vector transformed to E.coli strain DH5α and several procedures such as colony PCR, digestion and sequencing were assigned to confirm the consequence of the construct.Conclusions: The appearance of bacterial colonies on the plate containing different concentrations of streptomycin indicated the strength of resistance to streptomycin which showed the bacterial cells capability to express aadA gene under the controls of chloroplast regulatory elements.