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A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Molecular Genetics and Genomics".

Deadline for manuscript submissions: closed (31 July 2023) | Viewed by 26355

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Dr. Marsel Kabilov

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Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia
Interests: genome sequencing; next generation sequencing; metagenomics; metabarcoding, transcriptomics, translatomics

Special Issue Information

Dear Colleagues,

The advent of high-throughput sequencing platforms opened a new era in biology by allowing researchers to obtain and analyze huge data bodies. Every year, novel methods and techniques emerge which broaden our capabilities and interests, reaching far beyond the central dogma of molecular biology, i.e., DNA-seq, RNA-seq, and Ribo-seq. The transcriptomes of single cells (scRNA-seq), protein–nucleic interactions (Clip-seq), rare somatic mutations (DNA-seq with UMI), immunoglobulin repertoires, 3D arrangements of a genome (Hi-C), microbial communities, and many other approaches already acknowledge the presence of sequences. This has illuminated several conclusions about the molecular mechanisms of these processes. High-throughput sequencing is also sometimes limited by the imagination of researchers.

The first conference devoted to the high-throughput sequencing in genomics (HSG) took place in 2013. The third conference is scheduled for June 19–24, 2022, in Academgorodok (Novosibirsk, Russia). The HGS-2022 conference will feature talks about genomics of prokaryotes and eukaryotes, metagenomics of microbial communities, medical genomics, transcriptomics, protein–nucleic interactions, and translations, providing a forum to report and discuss novel high-throughput sequencing methodologies and their perspective application in research and technology.

Abstracts accepted for the conference can be published in a Special Issue of the journal IJMS (subject to peer review).

We look forward to receiving your contributions.

Dr. Marsel Kabilov
Guest Editor

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Keywords

metagenomics
genomics
metabarcoding
next generation sequencing
transcriptomics
translatomics
NGS data analysis

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22 pages, 3865 KiB

Open AccessArticle

Key FAD2, FAD3, and SAD Genes Involved in the Fatty Acid Synthesis in Flax Identified Based on Genomic and Transcriptomic Data

by Ekaterina M. Dvorianinova, Olga L. Zinovieva, Elena N. Pushkova, Daiana A. Zhernova, Tatiana A. Rozhmina, Liubov V. Povkhova, Roman O. Novakovskiy, Elizaveta A. Sigova, Anastasia A. Turba, Elena V. Borkhert, George S. Krasnov, Chengjiang Ruan, Alexey A. Dmitriev and Nataliya V. Melnikova

Int. J. Mol. Sci. 2023, 24(19), 14885; https://doi.org/10.3390/ijms241914885 - 4 Oct 2023

Cited by 4 | Viewed by 1782

Abstract

FAD (fatty acid desaturase) and SAD (stearoyl-ACP desaturase) genes play key roles in the synthesis of fatty acids (FA) and determination of oil composition in flax (Linum usitatissimum L.). We searched for FAD and SAD genes in the most widely used flax genome of the variety CDC Bethune and three available long-read assembled flax genomes—YY5, 3896, and Atlant. We identified fifteen FAD2, six FAD3, and four SAD genes. Of all the identified genes, 24 were present in duplicated pairs. In most cases, two genes from a pair differed by a significant number of gene-specific SNPs (single nucleotide polymorphisms) or even InDels (insertions/deletions), except for FAD2a-1 and FAD2a-2, where only seven SNPs distinguished these genes. Errors were detected in the FAD2a-1, FAD2a-2, FAD3c-1, and FAD3d-2 sequences in the CDC Bethune genome assembly but not in the long-read genome assemblies. Expression analysis of the available transcriptomic data for different flax organs/tissues revealed that FAD2a-1, FAD2a-2, FAD3a, FAD3b, SAD3-1, and SAD3-2 were specifically expressed in embryos/seeds/capsules and could play a crucial role in the synthesis of FA in flax seeds. In contrast, FAD2b-1, FAD2b-2, SAD2-1, and SAD2-2 were highly expressed in all analyzed organs/tissues and could be involved in FA synthesis in whole flax plants. FAD2c-2, FAD2d-1, FAD3c-1, FAD3c-2, FAD3d-1, FAD3d-2, SAD3-1, and SAD3-2 showed differential expression under stress conditions—Fusarium oxysporum infection and drought. The obtained results are essential for research on molecular mechanisms of fatty acid synthesis, FAD and SAD editing, and marker-assisted and genomic selection for breeding flax varieties with a determined fatty acid composition of oil. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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15 pages, 2323 KiB

Open AccessArticle

Comparative Transcriptomic Analysis of Three Common Liver Cell Lines

by Viktoriia Arzumanian, Mikhail Pyatnitskiy and Ekaterina Poverennaya

Int. J. Mol. Sci. 2023, 24(10), 8791; https://doi.org/10.3390/ijms24108791 - 15 May 2023

Cited by 2 | Viewed by 2340

Abstract

Background: Comparative transcriptomic analysis is a powerful approach for investigating the molecular mechanisms underlying various physiological and pathological processes, including liver disease. The liver is a vital organ with diverse functions, including metabolism and detoxification. In vitro models of liver cells, such as HepG2, Huh7, and Hep3B, have been widely used to study liver biology and pathology. However, there is limited information on the heterogeneity of these cell lines at the transcriptomic level. Objective: This study aimed to conduct a comparative transcriptomic analysis of three common liver cell lines (HepG2, Huh7, and Hep3B) using publicly available RNA-sequencing data. In addition, we compared these cell lines to primary hepatocytes, cells isolated directly from liver tissue and considered the gold standard for studying liver function and disease. Methods: Our study included sequencing data with the following criteria: total number of reads over 20,000,000, average read length of over 60 base pairs, Illumina sequencing, and non-treated cells. The data for the three cell lines were compiled: HepG2 (97 samples), Huh7 (39 samples), and Hep3B (16 samples). We performed differential gene expression analysis using the DESeq2 package, principal component analysis, hierarchical clustering on principal components, and correlation analysis to explore the heterogeneity within each cell line. Results: We identified numerous genes and pathways differentially expressed between HepG2, Huh7, and Hep3B, such as oxidative phosphorylation, cholesterol metabolism, and DNA damage. We report that the expression levels of important genes differ significantly between primary hepatocytes and liver cell lines. Conclusion: Our study provides new insights into the transcriptional heterogeneity of commonly used liver cell lines and highlights the importance of considering specific cell line. Consequently, transferring results without considering the heterogeneity of cell lines is impractical and may lead to inaccurate or distorted conclusions. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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13 pages, 2979 KiB

Open AccessArticle

Identification of Iguania Ancestral Syntenic Blocks and Putative Sex Chromosomes in the Veiled Chameleon (Chamaeleo calyptratus, Chamaeleonidae, Iguania)

by Katerina V. Tishakova, Dmitry Yu. Prokopov, Guzel I. Davletshina, Alexander V. Rumyantsev, Patricia C. M. O’Brien, Malcolm A. Ferguson-Smith, Massimo Giovannotti, Artem P. Lisachov and Vladimir A. Trifonov

Int. J. Mol. Sci. 2022, 23(24), 15838; https://doi.org/10.3390/ijms232415838 - 13 Dec 2022

Cited by 1 | Viewed by 2299

Abstract

The veiled chameleon (Chamaeleo calyptratus) is a typical member of the family Chamaeleonidae and a promising object for comparative cytogenetics and genomics. The karyotype of C. calyptratus differs from the putative ancestral chameleon karyotype (2n = 36) due to a smaller chromosome number (2n = 24) resulting from multiple chromosome fusions. The homomorphic sex chromosomes of an XX/XY system were described recently using male-specific RADseq markers. However, the chromosomal pair carrying these markers was not identified. Here we obtained chromosome-specific DNA libraries of C. calyptratus by chromosome flow sorting that were assigned by FISH and sequenced. Sequence comparison with three squamate reptiles reference genomes revealed the ancestral syntenic regions in the C. calyptratus chromosomes. We demonstrated that reducing the chromosome number in the C. calyptratus karyotype occurred through two fusions between microchromosomes and four fusions between micro-and macrochromosomes. PCR-assisted mapping of a previously described Y-specific marker indicates that chromosome 5 may be the sex chromosome pair. One of the chromosome 5 conserved synteny blocks shares homology with the ancestral pleurodont X chromosome, assuming parallelism in the evolution of sex chromosomes from two basal Iguania clades (pleurodonts and acrodonts). The comparative chromosome map produced here can serve as the foundation for future genome assembly of chameleons and vertebrate-wide comparative genomic studies. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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16 pages, 2321 KiB

Open AccessArticle

Genomic Diversity and Chromosomal Rearrangements in Neisseria gonorrhoeae and Neisseria meningitidis

by Boris Shaskolskiy, Dmitry Kravtsov, Ilya Kandinov, Ekaterina Dementieva and Dmitry Gryadunov

Int. J. Mol. Sci. 2022, 23(24), 15644; https://doi.org/10.3390/ijms232415644 - 9 Dec 2022

Cited by 2 | Viewed by 1964

Abstract

Chromosomal rearrangements in N. gonorrhoeae and N. meningitidis were studied with the determination of mobile elements and their role in rearrangements. The results of whole-genome sequencing and de novo genome assembly for 50 N. gonorrhoeae isolates collected in Russia were compared with 96 genomes of N. gonorrhoeae and 138 genomes of N. meningitidis from the databases. Rearrangement events with the determination of the coordinates of syntenic blocks were analyzed using the SibeliaZ software v.1.2.5, the minimum number of events that allow one genome to pass into another was calculated using the DCJ–indel model using the UniMoG program v.1.0. Population-level analysis revealed a stronger correlation between changes in the gene order and phylogenetic proximity for N. meningitidis in contrast to N. gonorrhoeae. Mobile elements were identified, including Correa elements; Spencer-Smith elements (in N. gonorrhoeae); Neisserial intergenic mosaic elements; IS elements of IS5, IS30, IS110, IS1595 groups; Nf1–Nf3 prophages; NgoФ1–NgoФ9 prophages; and Mu-like prophages Pnm1, Pnm2, MuMenB (in N. meningitidis). More than 44% of the observed rearrangements most likely occurred with the participation of mobile elements, including prophages. No differences were found between the Russian and global N. gonorrhoeae population both in terms of rearrangement events and in the number of transposable elements in genomes. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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15 pages, 2848 KiB

Open AccessArticle

Analysis of Expression Pattern of snoRNAs in Human Cells A549 Infected by Influenza A Virus

by Evgenii Zhuravlev, Mariia Sergeeva, Sergey Malanin, Rinat Amirkhanov, Dmitriy Semenov, Tatiana Grigoryeva, Andrey Komissarov and Grigory Stepanov

Int. J. Mol. Sci. 2022, 23(22), 13666; https://doi.org/10.3390/ijms232213666 - 8 Nov 2022

Cited by 4 | Viewed by 1940

Abstract

Small nucleolar RNAs (snoRNAs) are a highly expressed class of non-coding RNAs known for their role in guiding post-transcriptional modifications of ribosomal RNAs and small nuclear RNAs. Emerging studies suggest that snoRNAs are also implicated in regulating other vital cellular processes, such as pre-mRNA splicing and 3′-processing of mRNAs, and in the development of cancer and viral infections. There is an emerging body of evidence for specific snoRNA’s involvement in the optimal replication of RNA viruses. In order to investigate the expression pattern of snoRNAs during influenza A viral infection, we performed RNA sequencing analysis of the A549 human cell line infected by influenza virus A/Puerto Rico/8/1934 (H1N1). We identified 66 that were upregulated and 55 that were downregulated in response to influenza A virus infection. The increased expression of most C/D-box snoRNAs was associated with elevated levels of 5’- and 3’-short RNAs derived from this snoRNA. Analysis of the poly(A)+ RNA sequencing data indicated that most of the differentially expressed snoRNAs synthesis was not correlated with the corresponding host genes expression. Furthermore, influenza A viral infection led to an imbalance in the expression of genes responsible for C/D small nucleolar ribonucleoprotein particles’ biogenesis. In summary, our results indicate that the expression pattern of snoRNAs in A549 cells is significantly altered during influenza A viral infection. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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14 pages, 4231 KiB

Open AccessArticle

The Cytogenetic Map of the Nile Crocodile (Crocodylus niloticus, Crocodylidae, Reptilia) with Fluorescence In Situ Localization of Major Repetitive DNAs

by Svetlana A. Romanenko, Dmitry Yu. Prokopov, Anastasia A. Proskuryakova, Guzel I. Davletshina, Alexey E. Tupikin, Fumio Kasai, Malcolm A. Ferguson-Smith and Vladimir A. Trifonov

Int. J. Mol. Sci. 2022, 23(21), 13063; https://doi.org/10.3390/ijms232113063 - 27 Oct 2022

Cited by 4 | Viewed by 2872

Abstract

Tandemly arranged and dispersed repetitive DNA sequences are important structural and functional elements that make up a significant portion of vertebrate genomes. Using high throughput, low coverage whole genome sequencing followed by bioinformatics analysis, we have identified seven major tandem repetitive DNAs and two fragments of LTR retrotransposons in the genome of the Nile crocodile (Crocodylus niloticus, 2n = 32). The repeats showed great variability in structure, genomic organization, and chromosomal distribution as revealed by fluorescence in situ hybridization (FISH). We found that centromeric and pericentromeric heterochromatin of C. niloticus is composed of previously described in Crocodylus siamensis CSI-HindIII and CSI-DraI repetitive sequence families, a satellite revealed in Crocodylus porosus, and additionally contains at least three previously unannotated tandem repeats. Both LTR sequences identified here belong to the ERV1 family of endogenous retroviruses. Each pericentromeric region was characterized by a diverse set of repeats, with the exception of chromosome pair 4, in which we found only one type of satellite. Only a few repeats showed non-centromeric signals in addition to their centromeric localization. Mapping of 18S–28S ribosomal RNA genes and telomeric sequences (TTAGGG)_n did not demonstrate any co-localization of these sequences with revealed centromeric and pericentromeric heterochromatic blocks. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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9 pages, 1853 KiB

Open AccessArticle

Flow-Seq Evaluation of Translation Driven by a Set of Natural Escherichia coli 5′-UTR of Variable Length

by Ekaterina S. Komarova, Anna N. Slesarchuk, Maria P. Rubtsova, Ilya A. Osterman, Alexey E. Tupikin, Dmitry V. Pyshnyi, Olga A. Dontsova, Marsel R. Kabilov and Petr V. Sergiev

Int. J. Mol. Sci. 2022, 23(20), 12293; https://doi.org/10.3390/ijms232012293 - 14 Oct 2022

Cited by 1 | Viewed by 2206

Abstract

Flow-seq is a method that combines fluorescently activated cell sorting and next-generation sequencing to deduce a large amount of data about translation efficiency from a single experiment. Here, we constructed a library of fluorescent protein-based reporters preceded by a set of 648 natural 5′-untranslated regions (5′-UTRs) of Escherichia coli genes. Usually, Flow-seq libraries are constructed using uniform-length sequence elements, in contrast to natural situations, where functional elements are of heterogenous lengths. Here, we demonstrated that a 5′-UTR library of variable length could be created and analyzed with Flow-seq. In line with previous Flow-seq experiments with randomized 5′-UTRs, we observed the influence of an RNA secondary structure and Shine–Dalgarno sequences on translation efficiency; however, the variability of these parameters for natural 5′-UTRs in our library was smaller in comparison with randomized libraries. In line with this, we only observed a 30-fold difference in translation efficiency between the best and worst bins sorted with this factor. The results correlated with those obtained with ribosome profiling. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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19 pages, 2908 KiB

Open AccessArticle

The Structure of Stable Cellulolytic Consortia Isolated from Natural Lignocellulosic Substrates

by Grigory V. Gladkov, Anastasiia K. Kimeklis, Alexey M. Afonin, Tatiana O. Lisina, Olga V. Orlova, Tatiana S. Aksenova, Arina A. Kichko, Alexander G. Pinaev and Evgeny E. Andronov

Int. J. Mol. Sci. 2022, 23(18), 10779; https://doi.org/10.3390/ijms231810779 - 15 Sep 2022

Cited by 12 | Viewed by 2375

Abstract

Recycling plant matter is one of the challenges facing humanity today and depends on efficient lignocellulose degradation. Although many bacterial strains from natural substrates demonstrate cellulolytic activities, the CAZymes (Carbohydrate-Active enZYmes) responsible for these activities are very diverse and usually distributed among different bacteria in one habitat. Thus, using microbial consortia can be a solution to rapid and effective decomposition of plant biomass. Four cellulolytic consortia were isolated from enrichment cultures from composting natural lignocellulosic substrates—oat straw, pine sawdust, and birch leaf litter. Enrichment cultures facilitated growth of similar, but not identical cellulose-decomposing bacteria from different substrates. Major components in all consortia were from Proteobacteria, Actinobacteriota and Bacteroidota, but some were specific for different substrates—Verrucomicrobiota and Myxococcota from straw, Planctomycetota from sawdust and Firmicutes from leaf litter. While most members of the consortia were involved in the lignocellulose degradation, some demonstrated additional metabolic activities. Consortia did not differ in the composition of CAZymes genes, but rather in axillary functions, such as ABC-transporters and two-component systems, usually taxon-specific and associated with CAZymes. Our findings show that enrichment cultures can provide reproducible cellulolytic consortia from various lignocellulosic substrates, the stability of which is ensured by tight microbial relations between its components. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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18 pages, 2259 KiB

Open AccessArticle

Reorganization of the Landscape of Translated mRNAs in NSUN2-Deficient Cells and Specific Features of NSUN2 Target mRNAs

by Olga A. Kossinova, Alexander V. Gopanenko, Elena S. Babaylova, Alexey E. Tupikin, Marsel R. Kabilov, Alexey A. Malygin and Galina G. Karpova

Int. J. Mol. Sci. 2022, 23(17), 9740; https://doi.org/10.3390/ijms23179740 - 28 Aug 2022

Cited by 2 | Viewed by 2209

Abstract

The RNA cytosine C5 methyltransferase NSUN2 has a variety of RNA substrates and plays an important role in mRNA metabolism. NSUN2 binds to specific sequences enriched in exosomal mRNAs, suggesting its possible involvement in the sorting of mRNAs into exosomes. We applied the photoactivatable.4-thiouridine-enhanced cross-linking and immunoprecipitation assay involving high-throughput RNA sequencing (RNA-seq) to HEK293T cells to determine NSUN2 mRNA targets. NSUN2 cross-linking sites were found in more than one hundred relatively abundant mRNAs with a high GC content and a pronounced secondary structure. Then, utilizing RNA-seq for the total and polysome-associated mRNA from HEK293T cells with and without the knockdown of NSUN2, we identified differentially expressed genes, as well as genes with altered translational efficiency (GATEs). It turned out that the up-regulated GATE mRNAs were much shorter on average than the down-regulated ones, and their GC content was higher; moreover, they contained motifs with C residues located in GC-rich environments. Our findings reveal the specific features of mRNAs that make them potential targets for NSUN2 and expand our understanding of the role of NSUN2 in controlling translation and, possibly, in mRNA sorting into exosomes implemented through the methylation of cytosine residues. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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13 pages, 1383 KiB

Open AccessArticle

Nanopore Sequencing for De Novo Bacterial Genome Assembly and Search for Single-Nucleotide Polymorphism

by Maria G. Khrenova, Tatiana V. Panova, Vladimir A. Rodin, Maxim A. Kryakvin, Dmitrii A. Lukyanov, Ilya A. Osterman and Maria I. Zvereva

Int. J. Mol. Sci. 2022, 23(15), 8569; https://doi.org/10.3390/ijms23158569 - 2 Aug 2022

Cited by 9 | Viewed by 4769

Abstract

Nanopore sequencing (ONT) is a new and rapidly developing method for determining nucleotide sequences in DNA and RNA. It serves the ability to obtain long reads of thousands of nucleotides without assembly and amplification during sequencing compared to next-generation sequencing. Nanopore sequencing can help for determination of genetic changes leading to antibiotics resistance. This study presents the application of ONT technology in the assembly of an E. coli genome characterized by a deletion of the tolC gene and known single-nucleotide variations leading to antibiotic resistance, in the absence of a reference genome. We performed benchmark studies to determine minimum coverage depth to obtain a complete genome, depending on the quality of the ONT data. A comparison of existing programs was carried out. It was shown that the Flye program demonstrates plausible assembly results relative to others (Shasta, Canu, and Necat). The required coverage depth for successful assembly strongly depends on the size of reads. When using high-quality samples with an average read length of 8 Kbp or more, the coverage depth of 30× is sufficient to assemble the complete genome de novo and reliably determine single-nucleotide variations in it. For samples with shorter reads with mean lengths of 2 Kbp, a higher coverage depth of 50× is required. Avoiding of mechanical mixing is obligatory for samples preparation. Nanopore sequencing can be used alone to determine antibiotics-resistant genetic features of bacterial strains. Full article

(This article belongs to the Special Issue Selected Papers from the HSG-2022 Conference)

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