Morpho-histology of co-occurrence of somatic embryos, shoots, and inflorescences within a callus of Limonium 'Misty Blue'.

This is the first attempt to report the co-occurrence of somatic embryos, shoots, and inflorescences and their sequential development from stem cell niches of an individual callus mass through morpho-histological study of any angiosperm. In the presence of a proper auxin/cytokinin combination, precambial stem cells from the middle layer of a compact callus, which was derived from the thin cell layer of the inflorescence rachis of Limonium, expressed the highest level of totipotency and pluripotency and simultaneously developed somatic embryos, shoots, and inflorescences. This study also proposed the concept of programmed cell death during bipolar somatic embryo and unipolar shoot bud pattern formation. The unique feature of this research was the stepwise histological description of in vitro racemose inflorescence development. Remarkably, during the initiation of inflorescence development, either a unipolar structure with open vascular elements or an independent bipolar structure with closed vascular elements were observed. The protocol predicted the production of 6.6 ± 0.24 and 7.4 ± 0.24 somatic embryos and shoots, respectively, from 400 mg of callus, which again multiplied, rooted, and acclimatised. The plants' ploidy level and genetic fidelity were assessed randomly before acclimatisation by flow cytometry and inter simple sequence repeats (ISSR) marker analysis. Finally, the survivability and flower quality of the regenerated plants were evaluated in the field.

Direct Shoot From Root and True-to-Type Micropropagation of Limonium "Misty Blue" in Partially Immersed Culture on an Aluminum Mesh Raft.

Commercial plant tissue culture now primarily serves the ornamental horticulture industry. The main pillars of the commercial tissue culture business are scalability of production, cost reduction, limited labor involvement, high quality, and genetic homogeneity of propagated plants. Based on these requirements, the current protocol employs a partially immersed liquid culture medium supported by a flexible aluminum mesh raft with a wire stand to facilitate shoot organogenesis from the horizontally placed root explants and hold the plants upright for shoot multiplication and rooting of Limonium Misty Blue. It is a florist crop that is in high demand as both dried and fresh flower fillers in various floral decorations. The majority of cultivated Limonium or statice cultivars are heterozygous in nature and propagate commercially through in vitro propagation to cater to the huge demand for planting materials needed for flower production. This is the first protocol to describe direct shoot organogenesis from the roots in a liquid half-component of Murashige and Skoog's (1962) (MS) basal medium supplemented with 1.6 µM NAA and 1.1 µM BA. The regenerated shoots are multiplied and rooted at the same time on the raft in a MS-based liquid culture medium that included 0.44 µM BA and 1.07 µM NAA. In comparison to agar-gelled medium, plants cultured in liquid medium grow more quickly without any signs of hyperhydricity. In liquid medium, a clump of 4-5 shoots is formed from a single shoot explant within 4 weeks and are rooted simultaneously within 6 weeks. On average, seven explants may fit on each raft, so on average, 25 healthy plants are produced from a single bottle. The regenerated plants are easily hardened in the greenhouse, and using ISSR-based molecular markers, the genetic homogeneity of the randomly selected hardened plants can be determined.

Design modification of surgical drill bit for final osteotomy site preparation towards improved bone-implant contact.

Implant stability significantly impacts accelerated osseointegration, leading to faster patient recovery. Both primary and secondary stability necessitates superior bone-implant contact influenced by the surgical tool required to prepare the final osteotomy site. Besides, excessive shearing and frictional forces generate heat causing local tissue necrosis. Hence, surgical procedure necessitates proper irrigation with water to minimize heat generation. Notably, the water irrigation system removes bone chips and osseous coagulums, which may help accelerate osseointegration and improve bone-implant contact. The inferior bone-implant contact and thermal necrosis at the osteotomy site are primarily responsible for poor osseointegration and eventual failure. Therefore, optimizing tool geometry is key to minimizing shear force, heat generation, and necrosis during final osteotomy site preparation. The present study explores modified drilling tool geometry, especially cutting edge for osteotomy site preparation. The mathematical modeling is used to find out ideal cutting-edge geometry that facilitates drilling under relatively less operational force (0.55-5.24 N) and torque (98.8-154.5 N-mm) with a significant reduction (28.78%-30.87%) in heat generation. Twenty-three conceivable designs were obtained using the mathematical model; however, only three have shown promising results in static structural FEM platforms. These drill bits are designed for the final drilling operation and need to be carried out during the final osteotomy site preparation.

Factors influencing road traffic accidents causing maxillofacial injuries in Nalgonda District: prospective survey of 366 cases.

BACKGROUND: Road traffic accidents are the leading etiological factor for maxillofacial trauma in India. The incidence of these accidents is impacted by various cultural, socioeconomic, and behavioral factors the understanding of which is paramount in assessing their importance in influencing the incidence of maxillofacial injuries. METHODS: Data was collected via a questionnaire from 366 patients who reported with maxillofacial injuries due to Road Traffic Accidents to the casualty and maxillofacial OPD at a tertiary center in the Nalgonda District over a five-year period. Data collected included patient details, type of vehicle involved, speed of the vehicle, type of accident, location of the accident, the seating of the patient, presence of alcohol influence, usage of helmet or seatbelt and the injuries sustained by the patient. Statistical analysis was done using Chi squared test. RESULTS: 88.5%of the patients were male and 87.4% of the cases were injured in RTA involving two-wheeler vehicles. (50.3%) of the accidents took place between 6 pm to 12 am. 41.5% of cases reported their speed at the time of the accident as 40- 60 kmph. 42% of accidents were reported as skid accidents. 70.29% of accidents on rural roads occurred at night (between 6 pm and 6 am) as opposed to 29.71% during the day. Only 4.37% of cases reported wearing seatbelts or helmets. 51.17% of the participants who were driving reported or were observed as being under the influence of alcohol. CONCLUSIONS: The poor conditions of the roads, the lack of use of protective measures while driving, and the high incidence of driving under the influence of alcohol were seen to be the most significant contributing factors to road traffic accidents causing maxillofacial injuries in the Nalgonda population.

Somatic Embryogenesis of Anthurium andraeanum Linden., -A Tropical Florists' Plant.

The global floriculture market is expected to reach US$41.1 billion by 2027 at a CAGR of 5% over the analysis period 2020-2027; on the year 2020, the recorded market value in this trade was US$29.2 billion. The florists mainly use Anthurium andraeanum flowers in fashionable bouquets and floral arrangements because of their beautiful, attractive bright colored eye-catching spathe, candle-like spadix, prolonged vase life, etc. The cut flower industry always seeks elite cultivars and new hybrids of A. andraeanum, that in turn depend on the availability of large numbers of clonal planting propagules. In vitro somatic embryogenesis is an important technique for large-scale clonal propagation, development of transgenic plants, creation of new variety by somaclonal variation, mutagenesis on in vitro plants, and germplasm preservation for future use. Here, we describe the protocol of somatic embryogenesis of Anthurium andraeanum cv. Cancan, an important commercial cultivated variety. The protocol has been optimized by using 4 different types of culture media which are used during embryogenic callus induction, multiplication of callus, induction of somatic embryogenesis, and maturation plus conversion of embryos into plantlets. The protocol outlines the detailed methods from mother plant procurement to hardening of micro plants that is ready to transfer in the field and it can be used for large-scale commercial propagation.

Efficacy of Platelet-Rich Plasma Versus Hyaluronic Acid Following Arthrocentesis for Temporomandibular Joint Disc Disorders: A Randomized Controlled Trial.

Purpose: Injection of intra-articular medicaments is an accepted therapy for temporomandibular joint disorders (TMDs). This study compares the efficacy of arthrocentesis followed by platelet-rich plasma (PRP) as compared to hyaluronic acid (HA) injections for TMDs unresponsive to conservative therapy. PRP injection following arthrocentesis was hypothesized to be better compared to arthrocentesis alone or when combined with HA injection. Methods: Forty-seven patients with TMDs enrolled in an RCT were randomly assigned to three groups: Group A-PRP, Group B-HA or Group C-control group of arthrocentesis alone. Pre-operative evaluation and post-operative changes at 1-, 3-, 6-month intervals were assessed for improvement in pain, maximum mouth opening, joint sounds and excursive movements. Statistical significance was set at P value < 0.05. Results: Post-operative joint sounds were present in three out of 16 patients in Group A; six out of 15 patients in Group B and eight out of 16 patients in Group C at 6-month follow-up. For the remaining outcome variables, no statistical difference between groups was observed. Conclusion: Both medicaments produced significant clinical improvements when compared to the control group. When comparing PRP against HA, none showed superiority over the other.Clinical trial registration number: CTRI/2019/01/017076.

User-friendly tool kits for protein gel electrophoresis techniques: A training program for high school students.

Recent advancements in biochemical sciences have helped the researchers to explore the molecular logic of life inclusive of its multifarious expressions and explain many facts about the structure and functions of cellular macromolecules. Due to its simple and cost-effective nature, polyacrylamide gel electrophoresis (PAGE) has become the most favored technique for qualitative and quantitative examination of macromolecules. Major drawbacks of such modifications are the cost and operational complexities faced by naïve students. Many interlinking laboratory equipment are needed in the school laboratories for the conduct of even simple scientific experiment. Some of these costly modern equipment are inaccessible for students of small laboratories, and their alternatives are not easily available. Many of these laboratory equipment required for routine gel electrophoresis technique can be fabricated in their simplest form using off-the-shelf components. A short term biochemistry training program was executed for high school students to provide them "hands-on" training using newly modified equipment, which was proved to be an exciting way of learning biochemical gel separation techniques. © 2018 International Union of Biochemistry and Molecular Biology, 46(5):566-577, 2018.

A tetrad apparatus for protein gel casting, electrophoresis, staining, and scanning techniques with dual sensors for automatic detection of gel polymerization and protein migration.

Recent advancements in biochemical sciences have facilitated researchers to explore the structure and function of macro molecules in a cell. PAGE is one of the most favored and adapted laboratory techniques. Due to its simple and economical procedures, several variants or new modifications are routinely observed in the basic electrophoresis technique that comprises gel casting, electrophoresis, staining, and imaging process which consequently necessitates additional apparatuses/components in the laboratory. Operation of these additional apparatuses/components lengthens the pre- and postelectrophoresis procedures involving many intermittent tedious and time-consuming steps. A universal apparatus that can facilitate all such associated techniques is lacking and is of utmost importance for fast and effective results. An apparatus that can perform synchronized action of slab gel casting (16 × 16 cm), electrophoresis (SDS-PAGE), dye staining (Coomassie), and imaging (scanning) techniques with real-time monitoring through sensor technology is described in this article. The estimated cost (∼$150) of fabrication of the apparatus is very economical and simple assembly procedure of the main apparatus can be completed within ∼30 min after fabrication.

Regeneration of plantlets through somatic embryogenesis from root derived calli of Hibiscus sabdariffa L. (Roselle) and assessment of genetic stability by flow cytometry and ISSR analysis.

Induction of somatic embryogenesis and complete plantlet regeneration from callus culture of Hibiscus sabdariffa L. var. HS4288 has been made. Leaf and root explants were cultured on Murashige and Skoog (MS) and Driver-Kuniyuki Walnut (DKW) basal media supplemented with different concentrations of synthetic auxins and cytokinins. Root explants on DKW medium supplemented with 2.26µM 2, 4-Dichlorophenoxyacetic acid (2, 4-D) and 4.65µM kinetin (KIN) induced highest percentage (70%) of embryogenic calli. Average number of globular embryos per root derived callus produced within 6 weeks of culture initiation on MS media with different plant growth regulators (PGRs) ranged from 2.27±0.12 to 8.80±0.17 and that of cotyledonary embryos ranged from 0.00 to 2.53±0.20. On DKW medium comparatively more globular embryos (2.70±0.15 to 14.53±0.23) and cotyledonary embryos (0.00 to 8.90±0.17) were produced than that of MS medium. Regeneration of complete plantlets was highest (76.67%) when embryogenic calli with mature somatic embryos were grown on DKW medium containing 2.32µM KIN and 2.22µM 6-Benzyladenine (BA). Plants were primarily hardened in humidity, temperature and light controlled chamber and finally in a greenhouse showed 70% survival ability. Different stages of somatic embryogenesis process in the root derived embryogenic calli were elaborated in detail by morphological, histological and SEM study. The data were statistically analyzed by Duncan Multiple range test (p ≤ 0.05) and Principal component analysis (PCA). Flow cytometry and Inter-simple sequence repeats (ISSR) marker analysis confirmed that there was no genetic variation within the regenerated plants.

Simple and rapid system for two-dimensional gel electrophoresis technique: A laboratory exercise for high school students.

Polyacrylamide gel electrophoresis (PAGE) is the most classical technique favored worldwide for resolution of macromolecules in many biochemistry laboratories due to its incessant advanced developments and wide modifications. These ever-growing advancements in the basic laboratory equipments lead to emergence of many expensive, complex, and tricky laboratory equipments. Practical courses of biochemistry at high school or undergraduate levels are often affected by these complications. Two dimensional gel electrophoresis technique (2D-PAGE) used for resolving thousands of proteins in a gel is a combination of isoelectric focusing (first dimension gel electrophoresis technique) and sodium-dodecylsulphate PAGE (second dimension gel electrophoresis technique or SDS-PAGE). Two different laboratory equipments are needed to carry out effective 2D-PAGE technique, which also invites extra burden to the school laboratory. Here, we describe a low cost, time saving and simple gel cassette for protein 2D-PAGE technique that uses easily fabricated components and routine off-the-shelf materials. The performance of the apparatus was verified in a practical exercise by a group of high school students with positive outcomes. © 2018 by The International Union of Biochemistry and Molecular Biology, 46(3):237-244, 2018.

Multi-gel casting apparatus for vertical polyacrylamide gels with in-built solution flow system and liquid level detectors.

PAGE is the most widely used technique for the separation and biochemical analysis of biomolecules. The ever growing field of proteomics and genomics necessitates the analysis of many proteins and nucleic acid samples to understand further about the structure and function of cells. Simultaneous analysis of multiple protein samples often requires casting of many PAGE gels. Several variants of multi-gel casting/electrophoresis apparatuses are frequently used in research laboratories. Requirement of supplementary gels to match the growing demand for analyzing additional protein samples sometimes become a cause of concern. Available apparatuses are not amenable to and therefore, not recommended for any modification to accommodate additional gel casting units other than what is prescribed by the manufacturer. A novel apparatus is described here for casting multiple PAGE gels comprising four detachable components that provide enhanced practicability and performance of the apparatus. This newly modified apparatus promises to be a reliable source for making multiple gels in less time without hassle. Synchronized functioning of unique components broaden the possibilities of developing inexpensive, safe, and time-saving multi-gel casting apparatus. This apparatus can be easily fabricated and modified to accommodate desired number of gel casting units. The estimated cost (∼$300) for fabrication of the main apparatus is very competitive and effortless assembly procedure can be completed within ∼30 min.

A monophasic solution for isolation of RNA devoid of polymerase chain reaction-detectable genomic DNA contamination.

Commercially available reagents and published protocols are widely used for RNA isolation. However, genomic DNA contamination in isolated RNA is a potential problem. Here we describe a simple, inexpensive method for eliminating genomic DNA contamination beyond the level of PCR-based detection through reduction of the guanidine thiocyanate concentration (1.5M) in a single monophasic solution based on Chomczynski-Sacchi reagents. The new method can be used to isolate small and large RNA species of high quality and can be completed within an hour.

Identification and validation of a new male sex-specific ISSR marker in pointed gourd (Trichosanthes dioica Roxb.).

The aim of the present study was to develop a genetic sex marker for the pointed gourd (Trichosanthes dioica Roxb.) to allow gender determination at any stage in the life cycle. Screening of genomic DNA with intersimple sequence repeat (ISSR) primers was used to discover sex-specific touch-down polymerase chain reaction (Td-PCR) amplification products. Using pooled DNA from male and female genotypes and 42 ISSR primers, a putative male specific marker (~550 bp) was identified. DNA marker specific to male is an indication of existence of nonepigenetic factors involved in gender development in pointed gourd. The ISSR technique has proved to be a reliable technique in gender determination of pointed gourd genotypes at the seedling phenophase. The sex marker developed here could also be used as a starting material towards sequence characterization of sex linked genes for better understanding the developmental as well as evolutionary pathways in sexual dimorphism.