Multiple Sequence Research Articles

Fluorescent In Situ Hybridization Chain Reaction for RNA in the Drosophila Embryonic and Larval Central Nervous System.

In the Drosophila nerve cord, much is known about the generation of neurons from neuronal stem cells. Over the lifetime of a neuron, the cumulative expression of genes within that neuron determines its fate. Furthermore, gene expression in mature neurons determines their functional characteristics. It is therefore useful to visualize neural gene expression, which is often done via staining with antibodies to a protein of interest. In cases where there is no antibody to a desired gene product, or when it is useful to detect RNA rather than protein products, fluorescent in situ hybridization chain reaction for RNA (HCR RNA-FISH, or HCR for this protocol) can be used to detect and quantify RNA expression. RNA molecules reside predominantly in the cell soma, so HCR can facilitate determining neuron identity because somata position within the nerve cord is stereotyped across animals. HCR provides high-amplitude, high-fidelity signals. In principle, HCR can be broken down into a detection/hybridization stage and an amplification stage. During detection/hybridization, a probe set hybridizes to multiple sequences within a target gene. In the amplification step, concatemerized fluorescent hairpins bind to the hybridized probes. This two-step process increases the specificity of the fluorescent signal and helps reduce the likelihood of background fluorescence compared to traditional in situ hybridization techniques where the hybridizing probe itself contains the fluorescent signal. Here, we describe a protocol for using HCR to study gene expression in the Drosophila embryonic and larval nerve cord. We also describe how to combine HCR with immunofluorescence staining.

Guidelines in Designing a Universal Primer Mixture to Probe and Quantify Antibiotic-Resistant Genes Using the Polymerase Chain Reaction (PCR).

Multidrug resistance efflux pumps (MDREPs) in biofilm communities have become an increasingly expensive problem in clinical settings. Polymerase chain reaction (PCR)-based detection can be used to diagnose and characterize these genes, but this requires effective primer design to minimize false positives and negatives in test conclusions. A universal primer approach has previously been used to detect conserved core genes but not for accessory genes such as MDREPs. This study describes a guideline for the design of primers used in the detection of MDREP genes and an optimization approach for creating primers by using multiple sequence alignments to target conserved regions in silico, progressing from in silico to in vitro to generate working primers. Using thisapproach, this paper was able to generate primers to target sugE, a small multidrug resistance (SMR) protein found in microbial species. Primers were tested positively against synthetic DNA sequences but were inconsistent with DNA extracted from the organism of interest. Primer design informs the shortfalls of this detection technique and the difficulty in characterizing such genomic elements.

Chloroplast Genome Variation and Phylogenetic Relationships of Autochthonous Varieties of Vitis vinifera from the Don Valley.

The autochthonous grape varieties of the Don Valley, situated in southern Russia, constitute a distinctive element of regional cultural heritage. These varieties have been adapted over centuries to the region's specific local climatic and soil conditions. For the most part, these varieties are not imported from other countries. They are closely related to varieties found in Crimea and the North Caucasus. In this study, we obtained the first complete, unfragmented sequences of the chloroplast genomes of eight autochthonous varieties from the Don Valley and one from Crimea. We also performed a comparative analysis of their genomic features. The size of Vitis vinifera chloroplast genome sequences varied from 160,925 to 160,991 bp, depending on the cultivar, with a uniform GC ratio of 37.38%. Each genome consists of four subregions: a single copy region (LSC) ranging from 89,158 to 89,336 bp, a small single copy region (SSC) ranging from 19,070 to 19,073 bp, and a pair of inverted repeat regions (IRa and IRb) in the range of 26,292 to 26,353 bp. The chloroplast genomes of the studied V. vinifera varieties contained 130 genes, including 85 protein-coding genes, 8 rRNA genes, and 37 tRNA genes. The sequence divergence analysis has enabled the identification of four highly variable regions, which may be utilized as potential markers for phylogenetic analysis. The analysis revealed the presence of 58 to 61 SSRs and multiple long repeated sequences in the chloroplast genomes of these varieties. The phylogenetic analyses of the sequences obtained and complete chloroplast genomes available from public databases indicated that the majority of autochthonous V. vinifera varieties do not have a direct origin from any European variety.

Phylogenetic analysis and morphological characteristics of laccate Ganoderma specimens in Finland

ABSTRACT The Ganoderma lucidum complex includes fungi with similar morphologies but which are thought to represent different species. The lack of available type material and associated absence of multiple locus sequence data has complicated identification of these fungi. The aim of this study was to clarify the identity of the laccate Ganoderma species occurring in Finland by inferring a phylogeny using DNA sequences from available boreal-temperate material. DNA from Finnish isolates together with an older G. lucidum isolate originating from the United Kingdom was sequenced, and the morphological features of the Finnish specimens were examined. The phylogenetic analysis of the internal transcribed spacer region (ITS), the elongation factor 1-α (tef1), RNA polymerase II subunit (rpb2), and partial β-tubulin (β-tub) genes revealed that the G. lucidum isolate from the United Kingdom did not fall within a well-supported clade with other G. lucidum sequences or related species. The Finnish isolates were closely related to the G. tsugae lineage in tef1, rpb2, and β-tub phylogenies. However, G. tsugae appears morphologically distinct from the Finnish material. The results suggest that G. tsugae, or a species phylogenetically closely related to it, may occur in Finland. But further investigation into the relationship between G. tsugae and G. lucidum from Europe will be needed to clarify the identity of the laccate Ganoderma species in Finland.

Unraveling barley's PAL gene family: a genome-wide study on defense mechanisms against Puccinia graminis f. sp. tritici

Phenylalanine ammonia lyase (PAL) is a pivotal enzyme bridging primary and secondary phenylpropanoid metabolism, influencing plant growth, development, and stress responses. Despite extensive studies on PAL genes across various plant species, their investigation in barley, a critical staple food globally, has been relatively scarce. In this study, we have successfully identified 10 HvPAL genes, designated as HvPAL genes, in Hordeum vulgare (barley). These HvPAL genes were categorized based on their conserved sequences, which revealed patterns through MEME analysis and multiple sequence alignment. Interestingly, we found cis elements related to stress in the promoter regions of HvPAL genes, indicating their involvement in the response to pathogens. Furthermore, these gene promoters contained components associated with light, development, and hormone responsiveness. This suggests that they may play a role in hormonal developmental processes. MicroRNAs were also identified as regulators of the HvPAL genes we identified highlighting their significance in barley. To further investigate these gene expression patterns, we analyzed the RNA-seq data revealed the upregulating of HvPAL 2, HvPAL3, and HvPAL8, and downregulating HvPAL 5, HvPAL 6, and HvPAL9 genes in this study. This study focused on the regulation of PAL genes in response to 23 different races of Puccinia graminis f. sp. tritici in barley. These results suggest ways to improve traits and develop barley varieties that are resistant to pathogens by selectively increasing the expression of certain HvPAL genes that were not previously regulated. This thorough investigation aims to expand our knowledge of the versatility of the PAL gene family, providing insights for advancements in host -pathogen genetics.

Research on multi-sequence MR image synthesis CT algorithm based on unsupervised method

BackgroundUsing multi-sequence MRI to synthesize CT, combining different MRI sequences, and processing synthetic CT (sCT) based on unsupervised algorithms, to conduct more in-depth and comprehensive research on MR-based radiotherapy planning. MethodsFirst, we compare and analyze the effects of single-sequence MRI synthesis to CT to determine the quality of the synthesis and find a set of optimal sequence combinations. In addition, the study introduces a Staged Multi-Sequence Fusion Network (SMSF-Net) that employs an unsupervised approach to effectively fuse information from multiple MRI sequences. The staged sequence fusion module can efficiently extract information from different sequences and combine complementary information from different sequences. Finally, image encoding is processed using a multi-layer convolutional network to generate synthetic CT images. ResultsIn this study, the mean absolute error (MAE), root mean square error (RMSE), peak signal-to-noise ratio (PSNR), and structural similarity (SSIM) of the SMSF-Net are 18.85, 84.62, 34.31, and 0.962, which are superior to other single-sequence models and multi-sequence models. Experiments show that the improvement of SMSF-Net is statistically significant. In the qualitative comparison experiment, sCT synthesized by SMSF-Net is superior to other models in detail in the synthesis of complex areas of bones and nasal cavities. ConclusionThis study proposes a Staged SMSF-Net. As a multi-sequence MRI synthesis sCT algorithm based on unsupervised learning, SMSF-Net can use the complementary information between multiple sequences to improve the quality of synthetic sCT. This study has reference value for MRI-only radiotherapy planning.

Blocking uncertain mispriming errors of PCR

The polymerase chain reaction (PCR) plays a central role in genetic engineering and is routinely used in various applications, from biological and medical research to the diagnosis of viral infections. PCR is an extremely sensitive method for detecting target DNA sequences, but it is substantially error prone. In particular, the mishybridization of primers to contaminating sequences can result in false positives for virus tests. The blocker method, also called the clamping method, has been developed to suppress mishybridization errors. However, its application is limited by the requirement that the contaminating template sequence be known in advance. Here, we demonstrate that a mixture of multiple blocker sequences effectively suppresses the amplification of contaminating sequences even in the presence of uncertainty. The blocking effect was characterized by a simple model validated by experiments. Furthermore, the modeling allowed us to minimize the errors by optimizing the blocker concentrations. The results highlighted an inherent robustness of the blocker method in that fine-tuning the blocker concentrations is not necessary. Our method extends the applicability of PCR and other hybridization-based techniques, including genome editing, RNA interference, and DNA nanotechnology, by improving their fidelity.

Genome-wide identification of oxidosqualene cyclase genes regulating natural rubber in Taraxacum kok-saghyz.

Nine TkOSC genes have been identified by genome-wide screening. Among them, TkOSC4-6 might be more crucial for natural rubber biosynthesis in Taraxacum kok-saghyz roots. Taraxacum kok-saghyz Rodin (TKS) roots contain large amounts of natural rubber, inulin, and valuable metabolites. Oxidosqualene cyclase (OSC) is a key member for regulating natural rubber biosynthesis (NRB) via the triterpenoid biosynthesis pathway. To explore the functions of OSC on natural rubber producing in TKS, its gene family members were identified in TKS genome via genome-wide screening. Nine TkOSCs were identified, which were mainly distributed in the cytoplasm. Their family genes experienced a neutral selection during the evolution process. Overall sequence homology analysis OSC proteins revealed 80.23% similarity, indicating a highly degree of conservation. Pairwise comparisons showed a multiple sequence similarity ranging from 57% to 100%. Protein interaction prediction revealed that TkOSCs may interact with baruol synthase, sterol 1,4-demethylase, lupeol synthase and squalene epoxidase. Phylogenetic analysis showed that OSC family proteins belong to two branches. TkOSC promoter regions contain cis-acting elements related to plant growth, stress response, hormones response and light response. Protein accumulation analysis demonstrated that TkOSC4, TkOSC5 and TkOSC6 proteins had strong expression levels in the root, latex and plumular axis. Comparison of gene expression patterns showed TkOSC1, TkOSC4, TkOSC5, TkOSC6, TkOSC7, TkOSC8 and TkOSC9 might be important in regulating NRB. Combination of gene and protein results revealed TkOSC4-6 might be more crucial, and the data might contribute to a more profound understanding of the roles of OSCs for NRB in TKS roots.

AARS Online: A collaborative database on the structure, function, and evolution of the aminoacyl-tRNA synthetases.

The aminoacyl-tRNA synthetases (aaRS) are a large group of enzymes that implement the genetic code in all known biological systems. They attach amino acids to their cognate tRNAs, moonlight in various translational and non-translational activities beyond aminoacylation, and are linked to many genetic disorders. The aaRS have a subtle ontology characterized by structural and functional idiosyncrasies that vary from organism to organism, and protein to protein. Across the tree of life, the 22 coded amino acids are handled by 16 evolutionary families of Class I aaRS and 21 families of Class II aaRS. We introduce AARS Online, an interactive Wikipedia-like tool curated by an international consortium of field experts. This platform systematizes existing knowledge about the aaRS by showcasing a taxonomically diverse selection of aaRS sequences and structures. Through its graphical user interface, AARS Online facilitates a seamless exploration between protein sequence and structure, providing a friendly introduction to the material for non-experts and a useful resource for experts. Curated multiple sequence alignments can be extracted for downstream analyses. Accessible at www.aars.online, AARS Online is a free resource to delve into the world of the aaRS.

Magnetic resonance imaging of the dromedary camel carpus

BackgroundThe dromedary camel (Camelus dromedarius) carpal joint presents multiple joints and constitutes several bones and soft tissues. Radiography and/or ultrasonography of the carpus are challenging due to structural superimposition. High-field magnetic resonance imaging (MRI) technique precludes superimposed tissues and offers high soft tissue contrast in multiple sequences and planes. Hence, understanding the normal MRI anatomy is crucial during clinical investigations. Magnetic resonance imaging is highly sensitive for investigation of soft tissues and articular cartilage; therefore, it is extensively used for outlining joint anatomy and evaluation of a wide range of musculoskeletal conditions. MRI images of a specific anatomical region acquired by using multiple sequences in various planes are necessary for a complete MRI examination. Given the dearth of information on the MRI features of the dromedary camel carpus, the current study demonstrates the MRI appearance of the clinically significant structures in the camel carpus in various sequences and planes using a high-field 1.5 Tesla superconducting magnet. For this purpose, twelve cadaveric forelimbs, obtained from 6 clinically sound lameness free adult dromedary camels, were examined.ResultsThe cortex and medulla of the radius, carpal bones and metacarpus were evaluated. Articular cartilage of the carpal joints was depicted and showed intermediate intensity. Carpal tendons expressed lower signal intensity in all pulse sequences. The collateral and inter-carpal ligaments showed mixed signal intensity.ConclusionsThe obtained data outlines the validation of MRI for investigation of the camel carpus and could set as a reference for interpretation in clinical patients.

Engineering of Substrate-Binding Domain to Improve Catalytic Activity of Chondroitin B Lyase with Semi-Rational Design.

Dermatan sulfate and chondroitin sulfate are dietary supplements that can be utilized as prophylactics against thrombus formation. Low-molecular-weight dermatan sulfate (LMWDS) is particularly advantageous due to its high absorbability. The enzymatic synthesis of low-molecular-weight dermatan sulfates (LMWDSs) using chondroitin B lyase is a sustainable and environmentally friendly approach to manufacturing. However, the industrial application of chondroitin B lyases is severely hampered by their low catalytic activity. To improve the activity, a semi-rational design strategy of engineering the substrate-binding domain of chondroitin B lyase was performed based on the structure. The binding domain was subjected to screening of critical residues for modification using multiple sequence alignments and molecular docking. A total of thirteen single-point mutants were constructed and analyzed to assess their catalytic characteristics. Out of these, S90T, N103C, H134Y, and R159K exhibited noteworthy enhancements in activity. This study also examined combinatorial mutagenesis and found that the mutant H134Y/R159K exhibited a substantially enhanced catalytic activity of 1266.74 U/mg, which was 3.21-fold that of the wild-type one. Molecular docking revealed that the enhanced activity of the mutant could be attributed to the formation of new hydrogen bonds and hydrophobic interactions with the substrate as well as neighbor residues. The highly active mutant would benefit the utilization of chondroitin B lyase in pharmaceuticals and functional foods.

K3PO4–Mediated Synthesis of Chromeno–[3,4‐c]isoxazole from 3–Nitro–2H–chromene and α–Chloro Aldehyde via Michael Addition/C−Cl Bond Cleavage/Deformylation as Key Fragmentation Sequences

AbstractHere we report a transition metal– and oxidant–free annulation cascade method for the synthesis of chromeno–isoxazoles. The annulation reaction involves easily accessible starting materials, 3‐nitro‐2H‐chromene and α‐chloro aldehyde, yielding the structurally intriguing framework, chromeno[3,4‐c]isoxazoles employed by tripotassium phosphate as a base. The isoxazole ring construction over the 2H–chromene unit is believed to occur via a series of multiple reaction sequences. Here, the enolate generation/Michael addition/C−Cl bond cleavage/oxy–anionic addition to reactive tethered‐aldehyde/deformylation via retro–C−C and N−O bond cleavages are the most possible sequences which furnish the desired end–product in good chemical yields. The multiple reaction sequences occur in one‐pot at mild conditions, supported by experimental results and a few control experiments including DFT studies. This process exhibits a broad substrates scope illustrating good functional group compatibility towards both the nitrochromenes and aldehydes, and thus prove to be a general and efficient protocol for the synthesis of chromeno[3,4–c]isoxazoles. Overall, the use of nitrochromene as the limiting reagent, noteworthy functional group tolerance, scalability including great applications like chiral pool strategy towards the synthesis of enantioenriched chromeno‐isoxazole are the additional features.

Unveiling the genetic landscape of infectious laryngotracheitis virus in Switzerland: Evidence for vaccine-like and wild-type strains

Infectious laryngotracheitis (ILT) is a respiratory disease affecting chickens worldwide. Unlike many countries, Switzerland does not vaccinate against ILT. This study analysed ILT samples from 21 natural outbreaks in Switzerland using restriction fragment length polymorphism (RFLP) and multiple gene sequencing. Chicken embryo origin (CEO) and tissue culture origin (TCO) vaccine strains were included as references. Both vaccine strains were distinguishable, and 14 out of 21 samples resembled the CEO vaccine. Additionally, four distinct non-vaccine-like groups were identified. Sequencing of three genes from selected Swiss samples and those from neighbouring countries revealed four phylogenetic clades. Notably, four Swiss field strains formed two unique clades, not closely related to vaccine strains or ILTV from neighbouring countries. Overall, RFLP results were supported by sequencing data. This study demonstrates the presence of both vaccine-like and wild-type ILT viruses in Switzerland, where vaccination is de facto prohibited.

A putative scenario of how de novo protein-coding genes originate in the Saccharomyces cerevisiae lineage

BackgroundNovel protein-coding genes were considered to be born by re-organization of pre-existing genes, such as gene duplication and gene fusion. However, recent progress of genome research revealed that more protein-coding genes than expected were born de novo, that is, gene origination by accumulating mutations in non-genic DNA sequences. Nonetheless, the in-depth process (scenario) for de novo origination is not well understood.ResultsWe have conceived bioinformatic analysis for sketching a scenario for de novo origination of protein-coding genes. For each de novo protein-coding gene, we firstly identified an edge of a given phylogenetic tree where the gene was born based on parsimony. Then, from a multiple sequence alignment of the de novo gene and its orthologous regions, we constructed ancestral DNA sequences of the gene corresponding to both end nodes of the edge. We finally revealed statistical features observed in evolution between the two ancestral sequences. In the analysis of the Saccharomyces cerevisiae lineage, we have successfully sketched a putative scenario for de novo origination of protein-coding genes. (1) In the beginning was GC-rich genome regions. (2) Neutral mutations were accumulated in the regions. (3) ORFs were extended/combined, and then (4) translation signature (Kozak consensus sequence) was recruited. Interestingly, as the scenario progresses from (2) to (4), the specificity of mutations increases.ConclusionTo the best of our knowledge, this is the first report outlining a scenario of de novo origination of protein-coding genes. Our bioinformatic analysis can capture events that occur during a short evolutionary time by directly observing the evolution of the ancestral sequences from non-genic to genic. This property is suitable for the analysis of fast evolving de novo genes.

Assessing the role of evolutionary information for enhancing protein language model embeddings

Embeddings from protein Language Models (pLMs) are replacing evolutionary information from multiple sequence alignments (MSAs) as the most successful input for protein prediction. Is this because embeddings capture evolutionary information? We tested various approaches to explicitly incorporate evolutionary information into embeddings on various protein prediction tasks. While older pLMs (SeqVec, ProtBert) significantly improved through MSAs, the more recent pLM ProtT5 did not benefit. For most tasks, pLM-based outperformed MSA-based methods, and the combination of both even decreased performance for some (intrinsic disorder). We highlight the effectiveness of pLM-based methods and find limited benefits from integrating MSAs.

Metasurface Enabled Multi‐Target and Multi‐Wavelength Diffraction Neural Networks

AbstractBenefiting from low power consumption and high processing speed, there is a growing interest in diffraction neural networks (DNNs), which are typically showcased with 3D printing devices, leading to large volumes, high costs, and low levels of integration. Metasurfaces can desirably manipulate wavefronts of electromagnetic waves, providing a compact platform for mimicking DNNs with novel functions. Although multi‐wavelength and multi‐target recognition provides a richer and more detailed understanding of complex environments, existing architectures are primarily trained to classify a single target at a specific wavelength. A metasurface approach is proposed to design multiplexed DNNs that can classify multiple targets and spatial sequences across various wavelengths in multiple channels. To realize multi‐task processing, the dielectric metasurface is designed based on phase and wavelength multiplexing, which can integrate multi‐target DNNs with different tasks such as operating at distinct wavelengths and classifying diverse targets. The efficacy of this method is exemplified through the numerical simulation and experimental demonstration of recognizing a single target with two wavelengths, two targets at a single wavelength, and two targets at dual wavelengths. This compact metasurface approach enables the design of multi‐target and multi‐wavelength DNNs, opening a new window to develop massively parallel processing and versatile artificial intelligence systems.

Genomic profiling of pan-drug resistant proteus mirabilis Isolates reveals antimicrobial resistance and virulence gene landscape

Proteus mirabilis is a gram-negative pathogen that caused significant opportunistic infections. In this study we aimed to identify antimicrobial resistance (AMR) genes and virulence determinants in two pan-drug resistant isolate “Bacteria_11” and “Bacteria_27” using whole genome sequencing. Proteus mirabilis “Bacteria_11” and “Bacteria_27” were isolated from two different hospitalized patients in Egypt. Antimicrobial susceptibility determined using Vitek 2 system, then whole genome sequencing (WGS) using MinION nanopore sequencing was done. Antimicrobial resistant genes and virulence determinants were identified using ResFinder, CADR AMR database, Abricate tool and VF analyzer were used respectively. Multiple sequence alignment was performed using MAFFT and FastTree, respectively. All genes were present within bacterial chromosome and no plasmid was detected. “Bacteria_11” and “Bacteria_27” had sizes of approximately 4,128,657 bp and 4,120,646 bp respectively, with GC content of 39.15% and 39.09%. “Bacteria_11” and “Bacteria_27” harbored 43 and 42 antimicrobial resistance genes respectively with different resistance mechanisms, and up to 55 and 59 virulence genes respectively. Different resistance mechanisms were identified: antibiotic inactivation, antibiotic efflux, antibiotic target replacement, and antibiotic target change. We identified several genes associated with aminoglycoside resistance, sulfonamide resistance. trimethoprim resistance tetracycline resistance proteins. Also, those responsible for chloramphenicol resistance. For beta-lactam resistance, only blaVEB and blaCMY-2 genes were detected. Genome analysis revealed several virulence factors contribution in isolates pathogenicity and bacterial adaptation. As well as numerous typical secretion systems (TSSs) were present in the two isolates, including T6SS and T3SS. Whole genome sequencing of both isolates identify their genetic context of antimicrobial resistant genes and virulence determinants. This genomic analysis offers detailed representation of resistant mechanisms. Also, it clarifies P. mirabilis ability to acquire resistance and highlights the emergence of extensive drug resistant (XDR) and pan-drug resistant (PDR) strains. This may help in choosing the most appropriate antibiotic treatment and limiting broad spectrum antibiotic use.

Genetic heterogeneity of duck circovirus first detected in geese from China

Duck circovirus (DuCV) can infect domestic and wild ducks, retarding growth and suppressing immunity, thereby increasing the possibility of secondary infection by other pathogens. In this study, for the first time, 2 DuCV strains (G221116 and G210917) were identified in geese from China. To study the genetic characteristics of the 2 goose-originated DuCVs, multiple sequence alignment and phylogenetic analyses were perforemed according to genome sequences of 2 DuCV strains g and reference waterfowl circoviruses retrieved from the GenBank database. Pairwise analysis showed that the genome sequence identities between the 2 DuCVs with reference DuCV-1 and DuCV-2 strains were 80.95% to 98.24%, and 58.04% to 59.55% with Goose circovirus (GoCV). Phylogenetic analysis showed that the 2 DuCVs belonged to DuCV-1 and DuCV-2 genotypes. These results broaden our understanding of the genetic heterogeneity and evolution of DuCV and suggest trans-host transmission of DuCV between ducks and geese.

AlphaFold2 Predicts Alternative Conformation Populations in Green Fluorescent Protein Variants.

Artificial intelligence-based protein structure prediction methods such as AlphaFold2 have emerged as powerful tools for characterizing proteins sequence-structure relationship offering unprecedented opportunities for the molecular interpretation of biological and biochemical phenomena. While initially confined to providing a static representation of proteins through their global free-energy minimum, AlphaFold2 has demonstrated the ability to partially sample conformational landscapes, providing insights into protein dynamics, which is fundamental for interpreting and potentially tuning the function of natural and artificial proteins. In this study, we show that targeted column masking of AlphaFold2's multiple sequence alignment enables the characterization and estimation of the population ratio of the two main conformations of engineered green fluorescent proteins with alternative β-strands. The possibility of quickly estimating relative populations through AlphaFold2 predictions is expected to speed-up the computational design of related systems for sensing applications.

RFLP-kenzy: a new bioinformatics tool for in silico detection of key restriction enzyme in RFLP technique

BackgroundToday, several bioinformatics tools are available for analyzing restriction fragment length data. RFLP-kenzy is a new bioinformatic tool for identifying restriction key enzyme that cut at least 1 sequence and a maximum of n-1 sequence.ResultsThis bioinformatic tool helps researchers to select appropriate enzymes that yield different RFLP patterns, especially from overly identical sequences with single nucleotide mutation or other small variations. By using RFLP-kenzy, multiple DNA sequences could be analyzed simultaneously and the key enzymes list is provided. The present paper also demonstrates the ability of RFLP-kenzy to identify the key enzymes through the analysis of 16S rRNA sequences and the complete genome of various genera of microorganisms.ConclusionFrom the results, several key enzymes were provided indicating the importance of this new tool in the selection of appropriate restriction enzymes.