novel research question in molecular biology

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• Published: 02 March 2023

Methods in molecular biology and genetics: looking to the future

• Diego A. Forero 1 &
• Vaibhav Chand 2  

BMC Research Notes volume  16 , Article number:  26 ( 2023 ) Cite this article

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In recent decades, advances in methods in molecular biology and genetics have revolutionized multiple areas of the life and health sciences. However, there remains a global need for the development of more refined and effective methods across these fields of research. In this current Collection, we aim to showcase articles presenting novel molecular biology and genetics techniques developed by scientists from around the world.

A brief overview of the development of methods of molecular biology and genetics

Since ancient times, humankind has recognized the influence of heredity, based on familial resemblance, selective breeding of livestock, and climate-adapted crops. Prior to Gregor Johann Mendel’s work in the nineteenth century, there was no clear scientific theory to explain heredity. Mendel’s work remained essentially theoretical until the discovery of DNA and confirmation of its role as the principal agent of heredity in organisms in the twentieth century [ 1 ]. In addition, the resolution of the DNA structure paved the way for the invention of the Polymerase Chain Reaction (PCR) (by Kary Mullis), nucleotide synthesis [ 2 ] and the Sanger sequencing method [ 3 ] which revolutionized the field of genetics and led to the development of several sub-disciplines, including cytogenetics, biotechnology, bioprocess technology, and molecular biology. Automation of Sanger sequencing led to the Human Genome Project in 1990 [ 1 ], soon followed by sequencing the complete genomes of numerous other species of flora and fauna [ 4 ].

In recent decades, advances in methods in molecular biology and genetics have revolutionized multiple areas of life and health sciences [ 2 ]. As a major example from health sciences, PCR-based methods have advanced our understanding of the aetiology of a myriad of acute and chronic diseases, in addition to allowing the diagnosis of multiple disorders [ 1 , 5 ]. As a recent global application of molecular methods, the PCR-based approaches have led to the processing of hundreds of millions of samples for the analysis of the SARS-CoV-2 virus [ 6 ]. In addition, molecular methods have been key for the creation of multiple companies, products and jobs [ 7 ].

The development of sequencing technologies and their iterative improvements have been instrumental in advancing the understanding of DNA and RNA, their identification, association with various proteins, their covalent modifications, the function of the genes they carry, and the function of the non-coding portion of DNA and RNA in normal and diseased cells, in pathogenic bacteria and viruses, and in plants [ 8 , 9 ]. By producing RNA-based vaccines, we were able to combat the recent SARS-CoV2 pandemic. This was made possible by sequencing and in vitro nucleotide synthesis technologies [ 10 ].

Gene editing technologies, such as restriction endonuclease digestion, transcription activator-like effector nucleases (TALENs), and the clustered regularly interspaced short palindromic repeats (CRISPR-Cas) system, are an additional development in the field of molecular biology that has aided in the understanding of DNA and genes. There is optimism about the use of CRISPR-Cas9 technology in the treatment of a wide variety of diseases, such as cancer, blood-related diseases, hereditary blindness, cystic fibrosis, viral diseases, muscular dystrophy, and Huntington´s disease, due to its precision and its constant improvement, in comparison with other gene-editing technologies [ 15 ].

Need for novel methods in molecular biology and genetics

There is a global need for the development of novel methods for molecular biology and genetics. Particularly, in the area of human health, there is a need for further approaches that facilitate point-of-care molecular analysis (particularly miniaturized and portable platforms), for infectious and non-transmissible diseases [ 11 ], the development of more efficient methods for DNA sequencing [ 3 ], which facilitate cost-effective genome-wide analysis of patients, among others.

In addition, three key factors would also help push this field forward: additional research comparing the performance of different methods for molecular biology [ 12 ], the broader use of reporting standards (such as the Minimum Information for Publication of Quantitative Real-Time PCR Experiments -MIQE-, which describes details of experimental conditions) [ 13 ], and the increased participation of scientists from the Global South.

Although older techniques, such as x-ray crystallography, gene cloning, PCR, and sequencing, have been instrumental in the study of various aspects of genetics, these techniques have several limitations that result in gaps, missing links, and incomplete understanding of the genome. Advances in these techniques are needed to fill in these missing pieces of the puzzle to better comprehend genetics and accelerate the discovery of the causes of various genetically linkeddiseases. From a technological standpoint, the accuracy of sequencing and coverage across the genome remain major issues, especially for GC-rich regions and long homopolymer stretches of DNA. Furthermore, the short read lengths generated by the majority of current platforms severely restrict our ability to accurately characterize large repeat regions, numerous indels, and structural variation, rendering large portions of the genome opaque or inaccurate. Fragmentation of the genome for sequencing continues to be a major source of disruption in the continuity of the correct genomic sequence [ 14 , 15 ].

Recent advances in CRISPR technology provide hope for the medical treatment of cancer and other fatal diseases. Despite significant advances in this field, a number of technical obstacles remain, including off-target activity, insufficient indel or low homology-directed repair (HDR) efficiency, in vivo delivery of the Cas system components, and immune responses. This requires a substantial amount of technological advancement or the creation of new, superior methods to combat severe diseases with minimal side effects [ 14 , 16 ].

Additional considerations

As high-throughput, automated methods commonly produce very large amounts of data, deeper interaction between wet-lab and dry-lab researchers is required, to facilitate the design of efficient assays [ 17 ] and allow effective analysis and interpretation of results. Interdisciplinary collaborations, between biologists, engineers and professionals in the health sciences, might lead to newer and better methods of addressing current and future needs.

Further collaborations between scientists from academia and industry (in addition to researchers from government agencies) [ 18 ] would help to facilitate the development of novel methods, and aid in promoting their implementation around the world. For many countries, the main barrier to the broad use of molecular methods is the high cost of equipment and reagents [ 19 ]. Strategies aimed at lowering costs would be helpful for multiple institutions around the globe. In terms of intellectual property, fair licensing to institutions in the Global South as well as the implementation of Open Innovation and Open Science policies would be appropriate [ 20 ].

Overview of the current collection

In this current Collection, we are calling for articles showcasing novel methods from molecular biology and genetics, written by scientists from around the world. It is our goal to compile a set of articles that will help to address the challenges faced by the fields of molecular biology and genetics and broaden our understanding of genetic disorders and potential treatment strategies. We invite researchers working on such methods to consider submitting to our collection.

Data availability

Not applicable.

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Acknowledgements

DAF has been previously supported by research grants from Minciencias and Areandina. VC has been previously supported by research grants from NIH and VA.

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Diego A. Forero

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DAF and VC wrote an initial draft of the manuscript. All authors read and approved the final manuscript.

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DAF is a Senior Editorial Board Member of BMC Research Notes. VC is a Guest Editorial Board Member of BMC Research Notes.

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DAF is a medical doctor, Ph.D. in Biomedical Sciences and Professor and Research Leader at the School of Health and Sport Sciences, Fundación Universitaria del Área Andina (Bogotá, Colombia). He has worked with multiple methods of molecular biology and genetics and is an author of more than 100 articles in international journals, has been peer reviewer for more than 115 international scientific journals, in addition to being part of editorial boards of several international journals. VC is a Research Assistant Professor in the Department of Biochemistry and Molecular Genetics at the University of Illinois at Chicago. His expertise in Biochemistry, Molecular Biology, Genetics, Oncology, and Cancer Biology is extensive. He is an invited reviewer for more than fourteen international peer review journals and is the author of fourteen articles with high impact.

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Forero, D.A., Chand, V. Methods in molecular biology and genetics: looking to the future. BMC Res Notes 16 , 26 (2023). https://doi.org/10.1186/s13104-023-06298-y

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First step to untangle DNA: Supercoiled DNA captures gyrase like a lasso ropes cattle

Picture in your mind a traditional "landline" telephone with a coiled cord connecting the handset to the phone. The coiled telephone cord and the DNA double helix that stores the genetic material in every cell in the body have one thing in common; they both supercoil, or coil about themselves, and tangle in ways that can be difficult to undo. In the case of DNA, if this overwinding is not dealt with, essential processes such as copying DNA and cell division grind to a halt. Fortunately, cells have an ingenious solution to carefully regulate DNA supercoiling.

In this study published in the journal Science , researchers at Baylor College of Medicine, Université de Strasbourg, Université Paris Cité and collaborating institutions reveal how DNA gyrase resolves DNA entanglements. The findings not only provide novel insights into this fundamental biological mechanism but also have potential practical applications. Gyrases are biomedical targets for the treatment of bacterial infections and the similar human versions of the enzymes are targets for many anti-cancer drugs. Better understanding of how gyrases work at the molecular level can potentially improve clinical treatments.

Some DNA supercoiling is essential to make DNA accessible to allow the cell to read and make copies of the genetic information, but either too little or too much supercoiling is detrimental. For example, the act of copying and reading DNA overwinds it ahead of the enzymes that read and copy the genetic code, interrupting the process. It's long been known that DNA gyrase plays a role in untangling the overwinding, but the details were not clear.

DNA minicircles and advanced imaging techniques reveal first step to untangle DNA

"We typically picture DNA as the straight double helix structure, but inside cells, DNA exists in supercoiled loops. Understanding the molecular interactions between the supercoils and the enzymes that participate in DNA functions has been technically challenging, so we typically use linear DNA molecules instead of coiled DNA to study the interactions," said study author Dr. Lynn Zechiedrich, Kyle and Josephine Morrow Chair in Molecular Virology and Microbiology and professor of theVerna and Marrs McLean Department of Biochemistry and Molecular Pharmacology at Baylor College of Medicine. "One goal of our laboratory has been to study these interactions using a DNA structure that more closely mimics the actual supercoiled and looped DNA form present in living cells."

After years of work, the Zechiedrich lab has created small loops of supercoiled DNA. In essence, they took the familiar straight linear DNA double helix and twisted it in either direction once, twice, three times or more and connected the ends together to form a loop. Their previous study looking at the 3-D structures of the resulting supercoiled minicircles revealed that these loops form a variety of shapes that they hypothesized enzymes such as gyrase would recognize.

In the current study, their hypothesis was proven correct. The team of researchers combined their expertise to study the interactions of DNA gyrase with DNA minicircles using recent technology advances in electron cryomicroscopy, an imaging technique that produces high-resolution 3-D views of large molecules, and other technologies.

"My lab has long been interested in understanding how molecular nanomachines operate in the cell. We have been studying DNA gyrases, very large enzymes that regulate DNA supercoiling," said co-corresponding author Dr. Valérie Lamour, associate professor at the Institut de Génétique et de Biologie Moléculaire et Cellulaire, Université de Strasbourg. "Among other functions, supercoiling is the cell's way of confining about 2 meters (6.6 feet) of linear DNA into the microscopic nucleus of the cell."

As the DNA supercoils inside the nucleus, it twists and folds in different forms. Imagine twisting that telephone cord mentioned at the beginning, several times on itself. It will overwind and form a loop by crossing over DNA chains, tightening the structure.

"We found, just as we had hypothesized, that gyrase is attracted to the supercoiled minicircle and places itself in the inside of this supercoiled loop," said co-author, Dr. Jonathan Fogg, senior staff scientist of molecular virology and microbiology, and biochemistry and molecular pharmacology in the Zechiedrich lab.

"This is the first step of the mechanism that prompts the enzyme for resolving DNA entanglements," Lamour said.

"DNA gyrase, now surrounded by a tightly supercoiled loop, will cut one DNA helix in the loop, pass the other DNA helix through the cut in the other, and reseal the break, which relaxes the overwinding and eases the tangles, regulating DNA supercoiling to control DNA activity," Zechiedrich said. "Imagine watching the rodeo. Like roping cattle with a lasso, supercoiled looped DNA captures gyrase in the first step. Gyrase then cuts one double-helix of the DNA lasso and passes the other helix through the break to get free."

Co-corresponding author, Dr. Marc Nadal, professor at the École Normale in Paris confirmed the observation of the path of the DNA wrapped in the loop around gyrase using magnetic tweezers, a biophysical technique that allows to measure the deformation and fluctuations in the length of a single molecule of DNA. Observing a single molecule provides information that is often obscured when looking at thousands of molecules in traditional so-called "ensemble" experiments in a test tube.

Interestingly, the "DNA strand inversion model" for gyrase activity was proposed in 1979 by Drs. Patrick O. Brown and the late Nicholas R. Cozzarelli, also in a Science paper, well before researchers had access to supercoiled minicircles or the 3-D molecular structure of the enzyme. "It's especially meaningful to me that 45 years later, we finally provide experimental evidence supporting their hypothesis because Nick was my postdoctoral mentor," Zechiedrich said.

"This work opens a myriad of perspectives to study the mechanism of this conserved class of enzymes, which are of great clinical value," Lamour said.

"This work supports new ideas on how DNA activities are regulated. We propose that DNA is not a passive biomolecule acted upon by enzymes, but an active one that uses supercoiling, looping and 3-D shapes to direct accessibility of enzymes such as gyrase to specific DNA sequences in a variety of situations, which will likely impact cellular responses to antibiotics or other treatments," Fogg said.

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• Antiviral drug
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Materials provided by Baylor College of Medicine . Note: Content may be edited for style and length.

Journal Reference :

• Marlène Vayssières, Nils Marechal, Long Yun, Brian Lopez Duran, Naveen Kumar Murugasamy, Jonathan M. Fogg, Lynn Zechiedrich, Marc Nadal, Valérie Lamour. Structural basis of DNA crossover capture by Escherichia coli DNA gyrase . Science , 2024; 384 (6692): 227 DOI: 10.1126/science.adl5899

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Top 100 Latest Research Topics in Molecular Biology 2024 [Updated]

Table of Contents

Top 100 Latest Research Topics in Molecular Biology

So, are you thinking, what are some of the hottest research topics in molecular biology right now? Here is a list of current research topics in molecular biology:

• The part of non-histone proteins in chromosome building and function during mitosis.
• Restoration of the fission yeast Cdc42 cell-polarity component via the Sty1 p38 stress-activated protein kinase pathway.
• CRISPR and gene editing
• Gradient sensing: Engineering yeast love affair.
• How to make a static cytokinetic channel out of spreading excitable waves.
• Curvature-driven positioning of Turing patterns on phase-separating curved membranes.
• Activator-inhibitor coupling between Rho signaling and actin assembly makes the cell cortex an excitable medium.
• The meiotic spindle and chromosomes in oocytes.
• The molecular mechanisms underlying microtubule nucleation.
• The regulation of cell polarity, in a systems context, under both normal and stress conditions.
• Nuclear envelope transmembrane protein regulation of tissue specific genome group in differentiation and disease.
• Stem cell research
• What is the molecular basis for the establishment and maintenance of CENP-A nucleosomes at centromeres?
• How do the outer kinetochore microtubule binding components such as the Ska and Ndc80 complexes cooperate to facilitate spindle driven chromosome segregation?
• How CPC, a key player required for eliminating incorrect kinetochore-microtubule attachment is targeted to   the kinetochore? 
• Cystic fibrosis.
• Biochemical analysis of a restriction enzyme;
• Calculating the activity of alkaline phosphatase.
• Measuring the selection coefficient of yeast expressing Xenopus TFIIIA.
• Measuring plasmid loss in bacteria.
• Drug delivery
• Measuring the mutagenic effects of ethidium bromide.
• Measuring the mutagenic effects of UV light.
• Cancer cell research
• Measuring effect of plasmid size on transformation efficiency.
• Exploring bacterial transformation conditions.
• Using PCR to compare ribosomal RNA from Different Organisms.
• Chemo-enzymatic Synthesis and Potential Applications of Novel Heterobicyclic Alkaloids.
• Evolving an enzymatic toolbox to make complex modified peptides.
• Genome mining of novel bacterial carbazole natural products from new bacterial strains.
• Biological suggestions of genetic variation in rice for mycorrhizal colonization.
• Physiological and molecular characterization of altered root hydraulic properties in rice due to allelic variation or genetic manipulation of plasma membrane intrinsic proteins (PIPs, a class of aquaporins).
• RNA interference (RNAi)
• Determination of the most appropriate cell sources and harvest methods for in vitro culturing of pig meat.
• Monoclonal antibodies.
• Investigating the potential of novel mimetic molecules to modulate the microphage polarization.
• DNA synthesis
• Novel regulation of platelet-endothelial crosstalk in settings of vascular disease.
• Developing method to rationally design anti-cancer drug combinations.
• The role of non-histone proteins in chromosome structure and function during mitosis.
• Unloading of homologous recombination factors is required for restoring double-stranded DNA at damage repair loci.
• Aneuploidy as a mechanism of adaptation to telomerase insufficiency.
• When there is not enough telomerase: telomerase insufficiency and genome integrity.
• Proteolysis-dependent regulation of telomerase catalytic subunit.
• Molecular pathways that coordinate telomere maintenance and DNA repair machineries.
• Structural evidence for Scc4-dependent localization of cohesion loading.
• The kinetochore controls crossover recombination during meiosis.
• Quantitative cross-linking/mass- spectrometry reveals subtle protein conformational changes.
• Nuclear envelope transmembrane protein regulation of tissue specific genome organization in differentiation and disease
• Investigating the molecular mechanisms of parasite-host interaction between the green alga pluvialis and its parasitic fungus P. sedebokerense .
• Novel antimicrobial peptides from fish blood.
• Bacteriophages – Applications for Biocontrol.
• XenoImport – Towards expanding natural metabolism.
• Alternative splicing, intrinsically disordered regions and higher-order complex formation.
• Transcriptomics and structural bioinformatics of ion channels.
• 3 and its chaperones in the nervous system.
• Role of the SWI/SNF complex in the nervous system.
• Cytokines & growth factors.

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Novel Approaches to Study Metals in Molecular Biology

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Basics of Molecular Biology

State Key Laboratory of Space Medicine Fundamentals and Application, Astronaut Research and Training Center of China, Beijing, 100094 China

Dingsheng Zhao

Molecular biology is the study of biology on molecular level. The field overlaps with areas of biology and chemistry, particularly genetics and biochemistry. Molecular biology chiefly concerns itself with understanding the interactions between the various systems of a cell, including the interactions between DNA (deoxyribonucleic acid), RNA (Ribonucleic acid) and protein biosynthesis as well as learning how these interactions are regulated [1] .

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