Non-SELEX method for aptamer selection against ß-casomorphin-7 peptide.

The non-systematic evolution of ligands by the exponential enrichment (non-SELEX) method was used in the present study for the selection of ß-casomorphin-7 (BCM-7)-specific aptamers. These aptamers were tested to evaluate their ability to detect BCM-7 peptide in the human urine sample. The method did not employ aptamer amplification and counterselection as used in conventional SELEX but included a negative round of selection. The selection was performed in a single day, and after 5 rounds, a total of 16 numbers of aptamer were identified through Sanger sequencing. Newly selected aptamers named sequence ID no. 3 have performed better than other aptamers in detecting the BCM-7 peptide. Sequence ID no. 3 was also compared with previously selected aptamers through the SELEX method and its performance was found to be better than old aptamers. The sensing experiment was tried on different platforms from magnetic beads to the membrane. In each strategy, satisfactory results were obtained with aptamers that recognized BCM-7 spiked in a human urine sample at a very low amount. The non-SELEX method is an easy and time-saving process for aptamer selection. Selection of viable aptamers from a large pool of sequences for sensing experiments is a tedious job; however, an attempt has been made to select aptamers on the basis of In Silico (http://www.unafold.org/, https://bioinformatics.ramapo.edu/QGRS/index.php) information, observing DNA band intensity on agarose gel and colorimetric results obtained on magnetic beads and membrane. These aptamers have the potential in biosensor making for detecting BCM-7 peptide in urine samples of autistic patients.

Aptamers based sensing of pregnancy associated glycoproteins (PAG) of bovine for early pregnancy detection.

Tosyl activated magnetic beads were used for aptamer selection against PAG- 7 and 18 proteins of bovine origin. PAG proteins were immobilized on beads with further addition of biotin tagged aptamer library. The recognition of aptamers with PAG was identified by ST-HRP based approach which was colorimetric in nature. The selected aptamers were sequenced and at the same time several new aptamers were identified. Later M-fold structure and G-quadruplex score of aptamers were analyzed for their selection. Those aptamers having high G value and complex structure were chosen. In dot blot assay, aptamers recognized PAG protein in an animal after 42 days of artificial insemination which later given birth to a healthy calf. Further the cross reactivity with serum of 0th day animal (post AI) or with non pregnant animal serum was minimal. Aptamers have also shown interaction with PAG protein of buffalo origin. These selected aptamers have commercial application especially in development of biosensors for early detection of pregnancy in bovine.

International Depository Authority and its Role in Microorganism's Deposition.

After the World Trade Organization's agreement on Trade Related Aspect of Intellectual Property Right, patents have come into major play. Patenting of work related to live organisms that have medical, agricultural and other uses is always a tedious, complicated and controversial job. In view of this an agreement called as Budapest Treaty was passed in 1977 for deposition of microorganisms in culture collection centers for patent purpose. To make a culture collection center an IDA the culture center has to follow rules and regulations made in Budapest Treaty. Today several culture collection centers are working as International Depository Authority (IDA) in India and abroad that are storing microorganisms for patent purpose. India has two IDA units; one is Microbial Culture Collection in National Centre for Cell Science Pune, India, and second one is Microbial Type Culture Collection and Gene Bank at Institute of Microbial Technology Chandigarh, India. IDA units are important for valuable sample depositions, supply of cultures, characterization of samples and conservation of biological materials.

Aptamers in Therapeutics.

Aptamers are single strand DNA or RNA molecules, selected by an iterative process known as Systematic Evolution of Ligands by Exponential Enrichment (SELEX). Due to various advantages of aptamers such as high temperature stability, animal free, cost effective production and its high affinity and selectivity for its target make them attractive alternatives to monoclonal antibody for use in diagnostic and therapeutic purposes. Aptamer has been generated against vesicular endothelial growth factor 165 involved in age related macular degeneracy. Macugen was the first FDA approved aptamer based drug that was commercialized. Later other aptamers were also developed against blood clotting proteins, cancer proteins, antibody E, agents involved in diabetes nephropathy, autoantibodies involved in autoimmune disorders, etc. Aptamers have also been developed against viruses and could work with other antiviral agents in treating infections.

Aptamer-based sensing of ß-casomorphin-7.

ß-Casomorphin-7 (BCM-7), a seven amino acid peptide, is released during digestion of ß-casein A1 variant of milk which is speculated to be associated with certain diseases. Fifteen ssDNA aptamers having high affinity toward BCM-7 were identified from a 72 nt long random library after ten rounds of systematic evolution of ligands by exponential enrichment. Dissociation constant values of selected aptamers were in the range of 7.7-156.7 nM. Seq6 aptamer exhibited the lowest Kd value. Nine aptamers were evaluated for their binding toward BCM-7, BCM-9A1, and BCM-9A2 peptides, and binding was variable. SeqU5 exhibited the lowest binding with BCM-9A1 and BCM-9A2. Aptamer-coated gold nanoparticles (GNPs) resulted in color change of GNPs in the presence of BCM-7, thereby establishing recognition of BCM-7 by aptamers. The enzyme-linked aptamer-sorbent assay (ELASA) was evaluated as an assay of BCM-7 in biological fluids. BCM-7-peroxidase competed with BCM-7 in ELASA, performed with BCM-7 solution and BCM-7 spiked urine pretreated with urease, plasma, and ß-casein digest samples.

Torque Loss in En-Masse Retraction of Maxillary Anterior Teeth Using Miniimplants with Force Vectors at Different Levels: 3D FEM Study.

OBJECTIVE: This FEM study was conducted to quantify the amount of torque loss in maxillary anterior teeth by applying force vectors from different levels to the anterior retraction hook at various heights and comparing with that of molar anchorage system. MATERIALS AND METHODS: Five 3D FEM models were constructed with force vectors at different levels: HOT-High Orthodontic Traction (13.5mm from archwire) to ARH1- Anterior Retraction Hook (5mm), HOT to ARH2 (8mm), LOT- Low Orthodontic Traction (8 mm) to ARH1, LOT to ARH2 and from conventional molar hook to ARH1. Mini-implants were placed buccally between the roots of second premolar and first molar. Torque loss was calculated by measuring the displacement of the teeth at crown tip and root apex in two planes i.e. sagittal and vertical using Y and Z axis respectively in all the five models. The results were statistically analyzed by using Kruskal Wallis ANOVA and Mann-Whitney U-test. RESULTS: HOT to ARH1 showed that the anterior teeth moved bodily (p =0.5127), followed by molar hook - ARH1(p=0.0495*) which showed mild uncontrolled tipping. Whereas the HOT- ARH2, LOT - ARH1,and LOT - ARH2 models exhibited uncontrolled tipping with maximum torque loss in LOT - ARH1 (p=0.0001*). CONCLUSION: It can be concluded that bodily movement with very minimal torque loss was observed in HOT-ARH1 model whereas the maximum torque loss was recorded in LOT-ARH2 model. Conventional molar anchorage group showed uncontrolled tipping with some amount of extrusion and anchor loss of posteriors.

An overview of biomedical literature search on the World Wide Web in the third millennium.

Complete access to the existing pool of biomedical literature and the ability to "hit" upon the exact information of the relevant specialty are becoming essential elements of academic and clinical expertise. With the rapid expansion of the literature database, it is almost impossible to keep up to date with every innovation. Using the Internet, however, most people can freely access this literature at any time, from almost anywhere. This paper highlights the use of the Internet in obtaining valuable biomedical research information, which is mostly available from journals, databases, textbooks and e-journals in the form of web pages, text materials, images, and so on. The authors present an overview of web-based resources for biomedical researchers, providing information about Internet search engines (e.g., Google), web-based bibliographic databases (e.g., PubMed, IndMed) and how to use them, and other online biomedical resources that can assist clinicians in reaching well-informed clinical decisions.