Scholarly Works, Fralin Life Sciences Institute

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3,4-Dihydroxyphenylacetaldehyde synthase and cuticle formation in insects
Liao, Chenghong; Liang, Jing; Han, Qian; Li, Jianyong (2018-06-02)
Cuticle is the most important structure that protects mosquitoes and other insect species from adverse environmental conditions and infections of microorganism. The physiology and biochemistry of insect cuticle formation have been studied for many years and our understanding of cuticle formation and hardening has increased considerably. This is especially true for flexible cuticle. The recent discovery of a novel enzyme that catalyzes the production of 3,4-dihydroxyphenylacetaldehyde (DOPAL) in insects provides intriguing insights concerning the flexible cuticle formation in insects. For convenience, the enzyme that catalyzes the production DOPAL from L-dopa is named DOPAL synthase. In this mini-review, we summarize the biochemical pathways of cuticle formation and hardening in general and discuss DOPAL synthase-mediated protein crosslinking in insect flexible cuticle in particular.
Analysis of the Aedes albopictus C6/36 genome provides insight into cell line utility for viral propagation
Miller, Jason R.; Koren, Sergey; Dilley, Kari A.; Puri, Vinita; Brown, David M.; Harkins, Derel M.; Thibaud-Nissen, Françoise; Rosen, Benjamin D.; Xiao-Guang, Chen; Tu, Zhijian Jake; Sharakhov, Igor V.; Sharakhova, Maria V.; Sebra, R.; Stockwell, T. B.; Bergman, N. H.; Sutton, G. G.; Phillippi, A. M.; Pieemarini, P. M.; Shabman, R. S. (2018-03)
The 50-year old Aedes albopictus C6/36 cell line is a resource for the detection, amplification, and analysis of mosquito-borne viruses including Zika, dengue, and chikungunya. The cell line is derived from an unknown number of larvae from an unspecified strain of Aedes albopictus mosquitoes. Toward improved utility of the cell line for research in virus transmission, we present an annotated assembly of the C6/36 genome.
Association of microRNAs with Argonaute proteins in the malaria mosquito Anopheles gambiae after blood ingestion
Fu, X.; Dimopoulos, G.; Zhu, J. (2017)
The Beginning of the End: A Chromosomal Assembly of the New World Malaria Mosquito Ends with a Novel Telomere
Compton, Austin; Liang, Jiangtao; Chen, Chujia; Lukyanchikova, Varvara; Qi, Yumin; Potters, Mark B.; Settlage, Robert; Miller, Dustin; Deschamps, Stephane; Mao, Chunhong; Llaca, Victor; Sharakhov, Igor V.; Tu, Zhijian Jake (Genetics Society of America, 2020-10-01)
Chromosome level assemblies are accumulating in various taxonomic groups including mosquitoes. However, even in the few reference-quality mosquito assemblies, a significant portion of the heterochromatic regions including telomeres remain unresolved. Here we produce a de novo assembly of the New World malaria mosquito, Anopheles albimanus by integrating Oxford Nanopore sequencing, Illumina, Hi-C and optical mapping. This 172.6 Mbps female assembly, which we call AalbS3, is obtained by scaffolding polished large contigs (contig N50 = 13.7 Mbps) into three chromosomes. All chromosome arms end with telomeric repeats, which is the first in mosquito assemblies and represents a significant step toward the completion of a genome assembly. These telomeres consist of tandem repeats of a novel 30-32 bp Telomeric Repeat Unit (TRU) and are confirmed by analyzing the termini of long reads and through both chromosomal in situ hybridization and a Bal31 sensitivity assay. The AalbS3 assembly included previously uncharacterized centromeric and rDNA clusters and more than doubled the content of transposable elements and other repetitive sequences. This telomere-to-telomere assembly, although still containing gaps, represents a significant step toward resolving biologically important but previously hidden genomic components. The comparison of different scaffolding methods will also inform future efforts to obtain reference-quality genomes for other mosquito species.
Borrelia burgdorferi peptidoglycan is a persistent antigen in patients with Lyme arthritis
Jutras, Brandon L.; Lochhead, Robert B.; Kloos, Zachary A.; Biboy, Jacob; Strle, Klemen; Booth, Carmen J.; Govers, Sander K.; Gray, Joe; Schumann, Peter; Vollmer, Waldemar; Bockenstedt, Linda K.; Steere, Allen C.; Jacobs-Wagner, Christine (National Academy of Sciences, 2019-06-17)
Lyme disease is a multisystem disorder caused by the spirochete Borrelia burgdorferi. A common late-stage complication of this disease is oligoarticular arthritis, often involving the knee. In ∼10% of cases, arthritis persists after appropriate antibiotic treatment, leading to a proliferative synovitis typical of chronic inflammatory arthritides. Here, we provide evidence that peptidoglycan (PG), a major component of the B. burgdorferi cell envelope, may contribute to the development and persistence of Lyme arthritis (LA). We show that B. burgdorferi has a chemically atypical PG (PGBb) that is not recycled during cell-wall turnover. Instead, this pathogen sheds PGBb fragments into its environment during growth. Patients with LA mount a specific immunoglobulin G response against PGBb, which is significantly higher in the synovial fluid than in the serum of the same patient. We also detect PGBb in 94% of synovial fluid samples (32 of 34) from patients with LA, many of whom had undergone oral and intravenous antibiotic treatment. These same synovial fluid samples contain proinflammatory cytokines, similar to those produced by human peripheral blood mononuclear cells stimulated with PGBb. In addition, systemic administration of PGBb in BALB/c mice elicits acute arthritis. Altogether, our study identifies PGBb as a likely contributor to inflammatory responses in LA. Persistence of this antigen in the joint may contribute to synovitis after antibiotics eradicate the pathogen. Furthermore, our finding that B. burgdorferi sheds immunogenic PGBb fragments during growth suggests a potential role for PGBb in the immunopathogenesis of other Lyme disease manifestations.
Chromosome-level genome assemblies of the malaria vectors Anopheles coluzzii and Anopheles arabiensis
Zamyatin, Anton; Avdeyev, Pavel; Liang, Jiangtao; Sharma, Atashi; Chen, Chujia; Lukyanchikova, Varvara; Alexeev, Nikita; Tu, Zhijian Jake; Alekseyev, Max A.; Sharakhov, Igor V. (Oxford University Press, 2021-03-01)
Background: Anopheles coluzzii and Anopheles arabiensis belong to the Anopheles gambiae complex and are among the major malaria vectors in sub-Saharan Africa. However, chromosome-level reference genome assemblies are still lacking for these medically important mosquito species. Findings: In this study, we produced de novo chromosome-level genome assemblies for A. coluzzii and A. arabiensis using the long-read Oxford Nanopore sequencing technology and the Hi-C scaffolding approach. We obtained 273.4 and 256.8 Mb of the total assemblies for A. coluzzii and A. arabiensis, respectively. Each assembly consists of 3 chromosome-scale scaffolds (X, 2, 3), complete mitochondrion, and unordered contigs identified as autosomal pericentromeric DNA, X pericentromeric DNA, and Y sequences. Comparison of these assemblies with the existing assemblies for these species demonstrated that we obtained improved reference-quality genomes. The new assemblies allowed us to identify genomic coordinates for the breakpoint regions of fixed and polymorphic chromosomal inversions in A. coluzzii and A. arabiensis. Conclusion: The new chromosome-level assemblies will facilitate functional and population genomic studies in A. coluzzii and A. arabiensis. The presented assembly pipeline will accelerate progress toward creating high-quality genome references for other disease vectors.
Comparative Genomics and Proteomic Analysis of Assimilatory Sulfate Reduction Pathways in Anaerobic Methanotrophic Archaea
Yu, Hang; Susanti, Dwi; McGlynn, Shawn E.; Skennerton, Connor T.; Chourey, Karuna; Iyer, Ramsunder; Scheller, Silvan; Tavormina, Patricia L.; Hettich, Robert L.; Mukhopadhyay, Biswarup; Orphan, Victoria J. (Frontiers, 2018-12-03)
Sulfate is the predominant electron acceptor for anaerobic oxidation of methane (AOM) in marine sediments. This process is carried out by a syntrophic consortium of anaerobic methanotrophic archaea (ANME) and sulfate reducing bacteria (SRB) through an energy conservation mechanism that is still poorly understood. It was previously hypothesized that ANME alone could couple methane oxidation to dissimilatory sulfate reduction, but a genetic and biochemical basis for this proposal has not been identified. Using comparative genomic and phylogenetic analyses, we found the genetic capacity in ANME and related methanogenic archaea for sulfate reduction, including sulfate adenylyltransferase, APS kinase, APS/PAPS reductase and two different sulfite reductases. Based on characterized homologs and the lack of associated energy conserving complexes, the sulfate reduction pathways in ANME are likely used for assimilation but not dissimilation of sulfate. Environmental metaproteomic analysis confirmed the expression of 6 proteins in the sulfate assimilation pathway of ANME. The highest expressed proteins related to sulfate assimilation were two sulfite reductases, namely assimilatory-type low-molecular-weight sulfite reductase (alSir) and a divergent group of coenzyme F-420-dependent sulfite reductase (Group II Fsr). In methane seep sediment microcosm experiments, however, sulfite and zero-valent sulfur amendments were inhibitory to ANME-2a/2c while growth in their syntrophic SRB partner was not observed. Combined with our genomic and metaproteomic results, the passage of sulfur species by ANME as metabolic intermediates for their SRB partners is unlikely. Instead, our findings point to a possible niche for ANME to assimilate inorganic sulfur compounds more oxidized than sulfide in anoxic marine environments.
Comparative Metabolomics of Fruits and Leaves in a Hyperdiverse Lineage Suggests Fruits Are a Key Incubator of Phytochemical Diversification
Schneider, Gerald F.; Salazar, Diego; Hildreth, Sherry B.; Helm, Richard F.; Whitehead, Susan R. (Frontiers, 2021-08-30)
Interactions between plants and leaf herbivores have long been implicated as the major driver of plant secondary metabolite diversity. However, other plant-animal interactions, such as those between fruits and frugivores, may also be involved in phytochemical diversification. Using 12 species of Piper, we conducted untargeted metabolomics and molecular networking with extracts of fruits and leaves. We evaluated organ-specific secondary metabolite composition and compared multiple dimensions of phytochemical diversity across organs, including richness, structural complexity, and variability across samples at multiple scales within and across species. Plant organ identity, species identity, and the interaction between the two all significantly influenced secondary metabolite composition. Leaves and fruit shared a majority of compounds, but fruits contained more unique compounds and had higher total estimated chemical richness. While the relative levels of chemical richness and structural complexity across organs varied substantially across species, fruit diversity exceeded leaf diversity in more species than the reverse. Furthermore, the variance in chemical composition across samples was higher for fruits than leaves. By documenting a broad pattern of high phytochemical diversity in fruits relative to leaves, this study lays groundwork for incorporating fruit into a comprehensive and integrative understanding of the ecological and evolutionary factors shaping secondary metabolite composition at the whole-plant level.
Complete Dosage Compensation in Anopheles stephensi and the Evolution of Sex-Biased Genes in Mosquitoes
Jiang, Xiaofang; Biedler, James K.; Qi, Yumin; Hall, Andrew Brantley; Tu, Zhijian Jake (Oxford University Press, 2015-06-09)
Complete dosage compensation refers to hyperexpression of the entire X or Z chromosome in organisms with heterogametic sex chromosomes (XYmale or ZW female) in order to compensate for having only one copy of the X or Z chromosome. Recent analyses suggest that complete dosage compensation, as in Drosophila melanogaster, may not be the norm. There has been no systematic study focusing on dosage compensation in mosquitoes. However, analysis of dosage compensation in Anopheles mosquitoes provides opportunities for evolutionary insights, as theXchromosome of Anopheles and that of its Dipteran relative,D. melanogaster formed independently from the same ancestral chromosome. Furthermore, Culicinae mosquitoes, including the Aedes genus, have homomorphic sex-determining chromosomes, negating the need for dosage compensation. Thus, Culicinae genes provide a rare phylogenetic context to investigate dosage compensation in Anopheles mosquitoes. Here, we performed RNA-seq analysis ofmale andfemale samplesof theAsian malariamosquitoAnopheles stephensiandtheyellow fevermosquitoAedes aegypti.Autosomaland X-linked genes in An. stephensi showed very similar levels of expression in both males and females, indicating complete dosage compensation. The uniformity of average expression levels of autosomal and X-linked genes remained when An. stephensi gene expression was normalized by that of their Ae. aegypti orthologs, strengthening the finding of complete dosage compensation in Anopheles. In addition,we comparatively analyzed the differentially expressed genes between adultmales and adult females in both species, investigated sex-biased gene chromosomal distribution patterns in An. stephensi and provided three examples where gene duplications may have enabled the acquisition of sex-specific expression during mosquito evolution.
Complete Genome Sequence of Bordetella pertussis Pelita III, the Production Strain for an Indonesian Whole-Cell Pertussis Vaccine
Efendi, Yusuf Sofyan; Susanti, Dwi; Tritama, Erman; Pasier, Michelle; Putri, Gilang Nadia Niwan; Raharso, Sugeng; Iskandar; Aditiawati, Pingkan; Giri-Rachman, Ernawati; Mukhopadhyay, Biswarup; Purwantini, Endang (2017-04)
PT Bio Farma, the sole World Health Organization-approved Indonesian vaccine producer, manufactures a whole-cell whooping cough vaccine (wP) that, as part of a pentavalent diphtheria-tetanus-pertussis/hepatitis B/Haemophilus influenzae b (DTP/HB/Hib) vaccine, is used in Indonesia and many other countries. We report here the whole-genome sequence for Bordetella pertussis Pelita III, PT Bio Farma's wP production strain.
The complete genome sequence of Staphylothermus marinus reveals differences in sulfur metabolism among heterotrophic Crenarchaeota
Anderson, Iain J.; Dharmarajan, Lakshmi; Rodriguez, Jason; Hooper, Sean; Porat, Iris; Ulrich, Luke E.; Elkins, James G.; Mavromatis, Kostas; Sun, Hui; Land, Miriam; Lapidus, Alla; Lucas, Susan; Barry, Kerrie W.; Huber, Harald; Zhulin, Igor B.; Whitman, William B.; Mukhopadhyay, Biswarup; Woese, Carl; Bristow, James; Kyrpides, Nikos C. (2009-04-02)
Background Staphylothermus marinus is an anaerobic, sulfur-reducing peptide fermenter of the archaeal phylum Crenarchaeota. It is the third heterotrophic, obligate sulfur reducing crenarchaeote to be sequenced and provides an opportunity for comparative analysis of the three genomes. Results The 1.57 Mbp genome of the hyperthermophilic crenarchaeote Staphylothermus marinus has been completely sequenced. The main energy generating pathways likely involve 2-oxoacid:ferredoxin oxidoreductases and ADP-forming acetyl-CoA synthases. S. marinus possesses several enzymes not present in other crenarchaeotes including a sodium ion-translocating decarboxylase likely to be involved in amino acid degradation. S. marinus lacks sulfur-reducing enzymes present in the other two sulfur-reducing crenarchaeotes that have been sequenced - Thermofilum pendens and Hyperthermus butylicus. Instead it has three operons similar to the mbh and mbx operons of Pyrococcus furiosus, which may play a role in sulfur reduction and/or hydrogen production. The two marine organisms, S. marinus and H. butylicus, possess more sodium-dependent transporters than T. pendens and use symporters for potassium uptake while T. pendens uses an ATP-dependent potassium transporter. T. pendens has adapted to a nutrient-rich environment while H. butylicus is adapted to a nutrient-poor environment, and S. marinus lies between these two extremes. Conclusion The three heterotrophic sulfur-reducing crenarchaeotes have adapted to their habitats, terrestrial vs. marine, via their transporter content, and they have also adapted to environments with differing levels of nutrients. Despite the fact that they all use sulfur as an electron acceptor, they are likely to have different pathways for sulfur reduction.
Crystal Structures of TbCatB and Rhodesain, Potential Chemotherapeutic Targets and Major Cysteine Proteases of Trypanosoma brucei
Kerr, Iain D.; Wu, Peng; Marion-Tsukamaki, Rachel; Mackey, Zachary B.; Brinen, Linda S. (PLOS, 2010-06-01)
Background: Trypanosoma brucei is the etiological agent of Human African Trypanosomiasis, an endemic parasitic disease of sub-Saharan Africa. TbCatB and rhodesain are the sole Clan CA papain-like cysteine proteases produced by the parasite during infection of the mammalian host and are implicated in the progression of disease. Of considerable interest is the exploration of these two enzymes as targets for cysteine protease inhibitors that are effective against T. brucei. Methods and Findings: We have determined, by X-ray crystallography, the first reported structure of TbCatB in complex with the cathepsin B selective inhibitor CA074. In addition we report the structure of rhodesain in complex with the vinylsulfone K11002. Conclusions: The mature domain of our TbCatNCA074 structure contains unique features for a cathepsin B-like enzyme including an elongated N-terminus extending 16 residues past the predicted maturation cleavage site. N-terminal Edman sequencing reveals an even longer extension than is observed amongst the ordered portions of the crystal structure. The TbCatNCA074 structure confirms that the occluding loop, which is an essential part of the substrate-binding site, creates a larger prime side pocket in the active site cleft than is found in mammalian cathepsin B-small molecule structures. Our data further highlight enhanced flexibility in the occluding loop main chain and structural deviations from mammalian cathepsin B enzymes that may affect activity and inhibitor design. Comparisons with the rhodesainNK11002 structure highlight key differences that may impact the design of cysteine protease inhibitors as anti-trypanosomal drugs.
EGR1 recruits TET1 to shape the brain methylome during development and upon neuronal activity
Sun, Zhixiong; Xu, Xiguang; He, Jianlin; Murray, Alexander; Sun, Ming-an; Wei, Xiaoran; Wang, Xia; McCoig, Emmarose; Xie, Evan; Jiang, Xi; Li, Liwu; Zhu, Jinsong; Chen, Jianjun; Morozov, Alexei; Pickrell, Alicia M.; Theus, Michelle H.; Xie, Hehuang David (2019-08-29)
Life experience can leave lasting marks, such as epigenetic changes, in the brain. How life experience is translated into storable epigenetic information remains largely unknown. With unbiased data-driven approaches, we predicted that Egr1, a transcription factor important for memory formation, plays an essential role in brain epigenetic programming. We performed EGR1 ChIP-seq and validated thousands of EGR1 binding sites with methylation patterns established during postnatal brain development. More specifically, these EGR1 binding sites become hypomethylated in mature neurons but remain heavily methylated in glia. We further demonstrated that EGR1 recruits a DNA demethylase TET1 to remove the methylation marks and activate downstream genes. The frontal cortices from the knockout mice lacking Egr1 or Tet1 share strikingly similar profiles in both gene expression and DNA methylation. In summary, our study reveals EGR1 programs the brain methylome together with TET1 providing new insight into how life experience may shape the brain methylome.
Exploring Rain as Source of Biological Control Agents for Fire Blight on Apple
Llontop, Marco E. Mechan; Hurley, Kelly; Tian, Long; Galeano, Vivian A. Bernal; Wildschutte, Hans K.; Marine, Sasha C.; Yoder, Keith S.; Vinatzer, Boris A. (2020-02-14)
Poor survival on plants can limit the efficacy of Biological Control Agents (BCAs) in the field. Yet bacteria survive in the atmosphere, despite their exposure to high solar radiation and extreme temperatures. If conditions in the atmosphere are similar to, or more extreme than, the environmental conditions on the plant surface, then precipitation may serve as a reservoir of robust BCAs. To test this hypothesis, two hundred and fifty-four rain-borne isolates were screened for in vitro inhibition of Erwinia amylovora, the causal agent of fire blight, as well as of other plant pathogenic bacteria, fungi and oomycetes. Two isolates showed strong activity against E. amylovora and other plant pathogenic bacteria, while other isolates showed activity against fungal and oomycete pathogens. Survival assays suggested that the two isolates that inhibited E. amylovora were able to survive on apple blossoms and branches similarly to E. amylovora. Pathogen population size and associated fire blight symptoms were significantly reduced when detached apple blossoms were treated with the two isolates before pathogen inoculation, however, disease reduction on attached blossoms within an orchard was inconsistent. Using whole genome sequencing, the isolates were identified as Pantoea agglomerans and P. ananatis, respectively. A UV-mutagenesis screen pointed to a phenazine antibiotic D-alanylgriseoluteic acid synthesis gene cluster as being at the base of the antimicrobial activity of the P. agglomerans isolate. Our work reveals the potential of precipitation as an under-explored source of BCAs, whole genome sequencing as an effective approach to precisely identify BCAs, and UV-mutagenesis as a technically simple screen to investigate the genetic basis of BCAs. More field trials are needed to determine the efficacy of the identified BCAs in fire blight control.
Extracellular-signal regulated kinase 8 of Trypanosoma brucei uniquely phosphorylates its proliferating cell nuclear antigen homolog and reveals exploitable properties
Valenciano Murillo, Ana L.; Knudsen, Giselle M.; Mackey, Zachary B. (Taylor & Francis, 2016-01-01)
The Trypanosoma brucei subspecies T. brucei gambiense and T. brucei rhodesiense are vector-borne pathogens that cause sleeping sickness also known as Human African Trypanosomiasis (HAT), which is fatal if left untreated. The drugs that treat HAT are ineffective and cause toxic side effects. One strategy for identifying safer and more effective HAT drugs is to therapeutically exploit essential gene targets in T. brucei. Genes that make up a basic mitogen-activated protein kinase (MAPK) network are present in T. brucei. Tb927.10.5140 encodes an essential MAPK that is homologous to the human extracellular-signal regulated kinase 8 (HsERK8) which forms a tight complex with the replication factor proliferating cell nuclear antigen (PCNA) to stabilize intracellular PCNA levels. Here we demonstrate that (TbPCNA) is uniquely phosphorylated on serine (S) and threonine (T) residues in T. brucei and that TbERK8 phosphorylates TbPCNA at each of these residues. The ability of an ERK8 homolog to phosphorylate a PCNA homolog is a novel biochemical property that is first demonstrated here in T. brucei and may be unique to this pathogen. We demonstrate that the potent HsERK8 inhibitor Ro318220, has an IC50 for TbERK8 that is several hundred times higher than its reported IC50 for HsERK8. This indicated that the active sites of TbERK8 and HsERK8 can be selectively inhibited, which provides a rational basis for discovering inhibitors that specifically target this essential parasite MAPK to kill the parasite.
Fluorescence Polarization Binding Assay for Aspergillus fumigatus Virulence Factor UDP-Galactopyranose Mutase
Qi, Jun; Oppenheimer, Michelle; Sobrado, Pablo (Hindawi, 2011-08-21)
Aspergillus fumigatus is an opportunistic human pathogenic fungus responsible for deadly lung infections in immunocompromised individuals. Galactofuranose (Galf) residues are essential components of the cell wall and play an important role in A. fumigatus virulence. The flavoenzyme UDP-galactopyranose mutase (UGM) catalyzes the isomerization of UDP-galactopyranose to UDP-galactofuranose, the biosynthetic precursor of Galf. Thus, inhibitors of UGM that block the biosynthesis of Galf can lead to novel chemotherapeutics for treating A. fumigatus-related diseases. Here, we describe the synthesis of fluorescently labeled UDP analogs and the development of a fluorescence polarization (FP) binding assay for A. fumigatus UGM (AfUGM). High-affinity binding to AfUGM was only obtained with the chromophore TAMRA, linked to UDP by either 2 or 6 carbons with Kd values of 2.6 ± 0.2 μM and 3.0 ± 0.7 μM, respectively. These values were ~6 times lower than when UDP was linked to fluorescein. The FP assay was validated against several known ligands and displayed an excellent Z′ factor (0.79 ± 0.02) and good tolerance to dimethyl sulfoxide.
Functional analysis of the promoter of an early zygotic gene KLC2 in Aedes aegypti
Hu, Wanqi; Tu, Zhijian Jake (2018-12-24)
Background Aedes aegypti is an important mosquito vector that transmits arboviruses that cause devastating diseases including Zika, dengue fever, yellow fever and chikungunya. Improved understanding of gene regulation in the early development of Ae. aegypti will facilitate genetic studies and help the development of novel control strategies of this important disease vector. Results In this study, we demonstrated through transgenic assays that the promoter of an endogenous early zygotic gene KLC2 could drive gene expression in the syncytial blastoderm and early cellular blastoderm, which is a stage that the developing germline and the rest of embryo are accessible to genetic manipulation. An unexpected expression of the reporter gene in transgenic male testes was also observed. Further analysis confirmed the expression of the endogenous KLC2 in the testes, which was not detected in the previous RNA sequencing data. Conclusions Our finding provided a new promoter element that can be used in future genetic studies and applications in Ae. aegypti. Moreover, our transgenic reporter assays showed that cautions are needed when interpreting RNA sequencing data as transient or tissue-specific transcription may go undetected by RNAseq.
A Gene-Based Method for Cytogenetic Mapping of Repeat-Rich Mosquito Genomes
Masri, Reem A.; Karagodin, Dmitriy A.; Sharma, Atashi; Sharakhova, Maria V. (MDPI, 2021-02-06)
Long-read sequencing technologies have opened up new avenues of research on the mosquito genome biology, enabling scientists to better understand the remarkable abilities of vectors for transmitting pathogens. Although new genome mapping technologies such as Hi-C scaffolding and optical mapping may significantly improve the quality of genomes, only cytogenetic mapping, with the help of fluorescence in situ hybridization (FISH), connects genomic scaffolds to a particular chromosome and chromosome band. This mapping approach is important for creating and validating chromosome-scale genome assemblies for mosquitoes with repeat-rich genomes, which can potentially be misassembled. In this study, we describe a new gene-based physical mapping approach that was optimized using the newly assembled Aedes albopictus genome, which is enriched with transposable elements. To avoid amplification of the repetitive DNA, 15 protein-coding gene transcripts were used for the probe design. Instead of using genomic DNA, complementary DNA was utilized as a template for development of the PCR-amplified probes for FISH. All probes were successfully amplified and mapped to specific chromosome bands. The genome-unique probes allowed to perform unambiguous mapping of genomic scaffolds to chromosome regions. The method described in detail here can be used for physical genome mapping in other insects.
Genetic resources for advanced biofuel production described with the Gene Ontology
Torto-Alalibo, Trudy; Purwantini, Endang; Lomax, Jane; Setubal, João C.; Mukhopadhyay, Biswarup; Tyler, Brett M. (Frontiers, 2014-10-10)
Dramatic increases in research in the area of microbial biofuel production coupled with high-throughput data generation on bioenergy-related microbes has led to a deluge of information in the scientific literature and in databases. Consolidating this information and making it easily accessible requires a unified vocabulary. The Gene Ontology (GO) fulfills that requirement, as it is a well-developed structured vocabulary that describes the activities and locations of gene products in a consistent manner across all kingdoms of life. The Microbial ENergy processes Gene Ontology (http://www.mengo.biochem.vt.edu) project is extending the GO to include new terms to describe microbial processes of interest to bioenergy production. Our effort has added over 600 bioenergy related terms to the Gene Ontology. These terms will aid in the comprehensive annotation of gene products from diverse energy-related microbial genomes. An area of microbial energy research that has received a lot of attention is microbial production of advanced biofuels. These include alcohols such as butanol, isopropanol, isobutanol, and fuels derived from fatty acids, isoprenoids, and polyhydroxyalkanoates. These fuels are superior to first generation biofuels (ethanol and biodiesel esterified from vegetable oil or animal fat), can be generated from non-food feedstock sources, can be used as supplements or substitutes for gasoline, diesel and jet fuels, and can be stored and distributed using existing infrastructure. Here we review the roles of genes associated with synthesis of advanced biofuels, and at the same time introduce the use of the GO to describe the functions of these genes in a standardized way.
Genetic resources for methane production from biomass described with the Gene Ontology
Purwantini, E.; Torto-Alalibo, T.; Lomax, J.; Setubal, João C.; Tyler, B. M.; Mukhopadhyay, B. (Frontiers, 2014-12-03)

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