Swati Chaudhary

A bouquet of DNA structures: Emerging diversity

Structural polymorphism of DNA has constantly been evolving from the time of illustration of the double helical model of DNA by Watson and Crick. A variety of non-canonical DNA structures have constantly been documented across the globe. DNA attracted worldwide attention as a carrier of genetic information. In addition to the classical Watson–Crick duplex, DNA can actually adopt diverse structures during its active participation in cellular processes like replication, transcription, recombination and repair. Structures like hairpin, cruciform, triplex, G-triplex, quadruplex, i-motif and other alternative non-canonical DNA structures have been studied at length and have also shown their in vivo occurrence. This review mainly focuses on non-canonical structures adopted by DNA oligonucleotides which have certain prerequisites for their formation in terms of sequence, its length, number and orientation of strands along with varied solution conditions. This conformational polymorphism of DNA might be the basis of different functional properties of a specific set of DNA sequences, further giving some insights for various extremely complicated biological phenomena. Many of these structures have already shown their linkages with diseases like cancer and genetic disorders, hence making them an extremely striking target for structure-specific drug designing and therapeutic applications.

Molecular mapping of the grain iron and zinc concentration, protein content and thousand kernel weight in wheat (Triticum aestivum L.)

Genomic regions responsible for accumulation of grain iron concentration (Fe), grain zinc concentration (Zn), grain protein content (PC) and thousand kernel weight (TKW) were investigated in 286 recombinant inbred lines (RILs) derived from a cross between an old Indian wheat variety WH542 and a synthetic derivative (Triticum dicoccon PI94624/Aegilops squarrosa [409]//BCN). RILs were grown in six environments and evaluated for Fe, Zn, PC, and TKW. The population showed the continuous distribution for all the four traits, that for pooled Fe and PC was near normal, whereas, for pooled Zn, RILs exhibited positively skewed distribution. A genetic map spanning 2155.3cM was constructed using microsatellite markers covering the 21 chromosomes and used for QTL analysis. 16 quantitative trait loci (QTL) were identified in this study. Four QTLs (QGFe.iari-2A, QGFe.iari-5A, QGFe.iari-7A and QGFe.iari-7B) for Fe, five QTLs (QGZn.iari-2A, QGZn.iari-4A, QGZn.iari-5A, QGZn.iari-7A and QGZn.iari-7B) for Zn, two QTLs (QGpc.iari-2A and QGpc.iari-3A) for PC, and five QTLs (QTkw.iari-1A, QTkw.iari-2A, QTkw.iari-2B, QTkw.iari-5B and QTkw.iari-7A) for TKW were identified. The QTLs together explained 20.0%, 32.0%, 24.1% and 32.3% phenotypic variation, respectively, for Fe, Zn, PC and TKW. QGpc.iari-2A was consistently expressed in all the six environments, whereas, QGFe.iari-7B and QGZn.iari-2A were identified in two environments each apart from pooled mean. QTkw.iari-2A and QTkw.iari-7A, respectively, were identified in four and three environments apart from pooled mean. A common region in the interval of Xgwm359-Xwmc407 on chromosome 2A was associated with Fe, Zn, and PC. One more QTL for TKW was identified on chromosome 2A but in a different chromosomal region (Xgwm382-Xgwm359). Two more regions on 5A (Xgwm126-Xgwm595) and 7A (Xbarc49-Xwmc525) were found to be associated with both Fe and Zn. A QTL for TKW was identified (Xwmc525-Xbarc222) in a different chromosomal region on the same chromosome (7A). This reflects at least a partly common genetic basis for the four traits. It is concluded that fine mapping of the regions of the three chromosomes of A genome involved in determining the accumulation of Fe, Zn, PC, and TKW in this mapping population may be rewarding.

Structure-Specific Ligand Recognition of Multistranded DNA Structures.

Structural polymorphism is an extremely significant phenomenon of nucleic acids, in which DNA and RNA oligonucleotide sequences are able to adapt various canonical, alternative and multistranded structures. These alternative forms of DNA and RNA have an enormous potential of participating in various cellular processes by recognizing ligands such as proteins, drugs and metal ions in a sequence and structure-specific manner. Such DNA-ligand interactions prove to be highly beneficial when exploited for therapeutic purposes. Many of these DNA/ RNA structures recognizing drugs have already proved their potential as anticancer, antibacterial, anthelmintic and antiviral properties. Over the last 2-3 decades, many mechanisms of DNA-drug interactions have been documented, but still many other new mechanisms are being explored. Designing new drugs with improved efficacy and specificity is of prime concern for all researchers which not only deals with the experiments related to synthesizing drugs, but also takes care of searching novel routes or agents for administration or delivery of these therapeutic agents by increasing their nuclear and cellular uptake. This review aims at explaining the structural polymorphs/ multistranded DNA structures and their interactions with pharmaceutical drugs in a structure-specific manner, along with their modes of interactions and biological relevance. This detailed overview of multistranded DNA structures and interacting drugs might further facilitate our understanding about molecular targets and drug development in a more precise manner for the larger benefit of mankind.

Spectroscopic Studies of the Binding Interactions of PhenothiazininumDyes (Thionine Acetate, Azure A and Azure B) with Calf-thymus DNA

The double helical structure of DNA offers various binding sites for the interaction of ligands or proteins. Interactions using minor groove, major groove, and through intercalation are the major types of binding mechanisms of DNA-ligand interactions. The lowering in the absorption intensity along with bathochromic shift is the indication of intercalation binding mode of the dye into the base pairs of the DNA. In this study, the interaction of phenothiazine dyes with calf-thymus DNA (ctDNA) in physiological buffer (pH 7.4) was studied using UV-visible, fluorescence, circular dichroism (CD), and UV-thermal denaturation spectroscopy. The binding constants were calculated at different temperatures with the help of fluorescence spectroscopy. CD signals signify that B-form of DNA might become more compact, upon binding of the dyes. Also, induced circular dichroism is observed which confirms the dye-DNA complex formation. Stabilization of DNA double helix upon binding with dyes was confirmed by the increase in Tm of ctDNA. Based on thermal melting profiles, it was found that thionine acetate is most promising in stabilizing the DNA double helix, in comparison to other two dyes. Also, binding constants calculated by fluorescence is in accordance with the thermal melting analysis. These results are indicative of the intercalation binding mode between dyes and the DNA. The binding affinity of the dyes to DNA is found to be in order as thionine acetate > azure A > azure B. Such preliminary studies facilitate our understanding about various types of DNAligand interactions and provide clues for designing new and more effective drugs.

Formation of G-wires, bimolecular and tetramolecular quadruplex: Cation-induced structural polymorphs of G-rich DNA sequence of human SYTX gene

An exceptional property of auto‐folding into a range of intra‐ as well as intermolecular quadruplexes by guanine‐rich oligomers (GROs) of promoters, telomeres and various other genomic locations is still one of the most attractive areas of research at present times. The main reason for this attention is due to their established in vivo existence and biological relevance. Herein, the structural status of a 20‐nt long G‐rich sequence with two G5 stretches (SG20) is investigated using various biophysical and biochemical techniques. Bioinformatics analysis suggested the presence of a 17‐nt stretch of this SG20 sequence in the intronic region of human SYTX (Synaptotagmin 10) gene. The SYTX gene helps in sensing out the Ca2+ ion, causing its intake in the pre‐synaptic neuron. A range of various topologies like bimolecular, tetramolecular and guanine‐wires (nano‐wires) was exhibited by the studied sequence, as a function of cations (Na+/K+) concentration. UV‐thermal denaturation, gel electrophoresis, and circular dichroism (CD) spectroscopy showed correlations and established a cation‐dependent structural switch. The G‐wire formation, in the presence of K+, may further be explored for its possible relevance in nano‐biotechnological applications.

Structural Switch from Hairpin to Duplex/Antiparallel G-Quadruplex at Single-Nucleotide Polymorphism (SNP) Site of Human Apolipoprotein E (APOE) Gene Coding Region

A gradual dementia, which leads to the loss of memory and intellectual abilities, is the main characteristics of Alzheimer’s disease. Amyloid-β (Aβ) plaques are the main components that accumulate and form clumps in the brains of people suffering from Alzheimer’s disease. Apolipoprotein E (APOE), an amyloid-binding protein is considered as one of the main genetic risk factor of the late-onset Alzheimer’s disease. Different isoforms of APOE gene named APOE2, APOE3, and APOE4 are known to exist, which differ from each other at certain positions involving single-nucleotide polymorphisms (SNPs). Out of these isoforms, APOE4 increases the risk of developing late-onset Alzheimer’s disease, whereas APOE3 is the most common among the general population. APOE4 differs from the common APOE3 by only one nucleotide at position +2985 (T to C), which results in immense alteration in the structure and function of the APOE gene. A combination of gel electrophoresis (polyacrylamide gel electrophoresis, PAGE), circular dichroism (CD), CD melting, thermal difference spectra and UV-thermal denaturation (TM) techniques was used to investigate the structural polymorphism associated with T → C single-nucleotide polymorphism (SNP) at the GC-rich sequence (d-TGGAGGACGTGTGCGGCCGCCT; APOE22T). Herein, we report that APOE22T DNA sequence switches between hairpin to antiparallel quadruplex from low to high oligomer concentration. On the contrary, its C-counterpart (APOE22C) forms hairpin as well as intermolecular antiparallel duplex structure. This structural change may possibly contribute to the protein recognition pattern, which, in turn, might control the APOE gene expression.

DNA Fingerprints: Advances in their Forensic Analysis Using Nanotechnology

For deciphering the secrets of forensic science, nanotechnology has quite extensively been utilized. Generally, for identifying the fingerprints, a lot of combination of different materials and film assemblies have already been utilized. Since the mode of interaction between the nanoparticles and fingerprint marks, is still not clearly understood, fabricating the nanoparticle assemblies for their identification is quite challenging. Complete identification of fingerprint marks which are generally because of a combination of some kind of proteins and fatty acids, is still a difficult task and is only partially being done with the help of different techniques. Nanotechnology has already shown immense potential in many fields like medicine, molecular biology, genetics, material science etc. and it has also marked promising potential even in forensic analysis studies. This review aims to discuss the details of the process of fingerprint formation, their role in forensic analysis along with the latest advances in the field of nanotechnology for their identification. This information may enhance our understanding about the progress in the forensic analysis of fingerprints, which may further be utilized in solving the puzzle of various criminal cases.

Genetic Variations: Heroes or Villains

Gene pool of every organism has shown a wide occurrence of genetic variations. Variations have not only been associated with diseases like cancer, turner syndrome, sickle cell anaemia, cystic fibrosis etc., but some of them have even been proved to be beneficial in certain cases like for increasing bone density, lowering down cholesterol level, and for developing malaria resistance. Genetic variations or switches have also been explored for their significant role in evolution of human species. Monogenic diseases are the inherited disorders caused by the mutations in a single gene. Single Nucleotide Polymorphism, structural variants and genomic rearrangements are considered as some of the forms of genetic mutations. Genetic mutations are the fundamental cause of some monogenic disorders such as Schizophrenia, Alzheimer’s disease, Hutchinson-Gilford Progeria syndrome (HGPS), Proteus syndrome and Congenital generalized hypertrichosis (CGH). However, some variants have even offered protection against a disease like blocking of HIV infection due to mutations in co-receptors needed for entry of infection into the cells. This review aims to provide insights into the role of variants present in the genes associated with the monogenic disorders in order to determine the underlying mechanism of the disease, which might further pave a way for the scientists to discover the therapeutic approaches for dealing with the same.

Genomic Databases and Softwares: In Pursuit of Biological Relevancethrough Bioinformatics

With the completion of human genome project, a plethora of information had been available for exploring various unanswered questions related to cell and molecular biology. Bioinformatics has been instrumental in unravelling the genetic, phenotypic, structural and functional aspects of the whole genome by using this information. Genomics and proteomics have become one of the most relevant fields after the advancements in computational analysis, interpretation, and modelling software. It can not only quite categorically describe the position of nucleotides and amino acids throughout genomes and proteomes respectively, but it also helps in performing the phylogenetic analysis, search for associated transcription factors, multiple sequence alignments, and many other relevant explorations/hunts. The advances in the knowledge of genetics acquired from molecular biology and bioinformatics are applied and point towards potential therapeutic strategies such as genome editing. This review has an aim of discussing some of the bioinformatics databases and software, which has been utilized for exploring the position of a DNA sequence, any associated single nucleotide polymorphism (SNP) related to a disease, on or nearby situated transcription factor binding sites followed by multiple sequence alignment of this sequence with other organisms. This study provides the insights in to the functional elements of any DNA, RNA or Protein sequence prior to exploring the structural polymorphism, which may regulate the gene expression. Also, this review briefly discusses the tools used for programmable nuclease–based genome editing technology.

Advancements in Characterization Techniques of Biopolymers: CyclicVoltammetry, Gel Electrophoresis, Circular Dichroism and FluorescenceSpectroscopy

Biopolymers such as DNA, RNA and proteins are the key players in the cellular processes like cell differentiation, cell growth, maintenance, repair, recombination, transcription, translation, etc. Immense efforts have been made for their isolation, purification, quantification and structural as well as functional characterization. Profound insights into the structural and functional characterization of these biopolymers could help us understand the intricate cellular machinery. A wide range of biochemical, biophysical, electrochemical and molecular biology techniques have really been beneficial in exploring the key and interdependent relationships between the structure and function of these biopolymers. Each instrumentation technique has its own advantage and disadvantage in terms of their applications, selectivity and sensitivity. There had been a lot of advancements in these techniques for achieving their desired goals, but still, lots of limitations need our attention and further improvements. For understanding, the extremely complicated biological machinery at molecular and cellular level, every small fragment of a cell has to be studied separately with a lot of intricacies. In this review, we present a brief update of the advancements made in the large repertoire of Cyclic Voltammetry (CV), Gel electrophoresis, Circular Dichroism (CD) and Fluorescence spectroscopy techniques which might be relevant for updating our knowledge of the pre-existing and well-established tools utilized by the scientists across the world.