Validation of Golden Gate Assemblies Using Highly Multiplexed Nanopore Amplicon Sequencing.

Golden Gate cloning has revolutionized synthetic biology. Its concept of modular, highly characterized libraries of parts that can be combined into higher order assemblies allows engineering principles to be applied to biological systems. The basic parts, typically stored in Level 0 plasmids, are sequence validated by the method of choice and can be combined into higher order assemblies on demand. Higher order assemblies are typically transcriptional units, and multiple transcriptional units can be assembled into multi-gene constructs. Higher order Golden Gate assembly based on defined and validated parts usually does not introduce sequence changes. Therefore, simple validation of the assemblies, e.g., by colony polymerase chain reaction (PCR) or restriction digest pattern analysis is sufficient. However, in many experimental setups, researchers do not use defined parts, but rather part libraries, resulting in assemblies of high combinatorial complexity where sequencing again becomes mandatory. Here, we present a detailed protocol for the use of a highly multiplexed dual barcode amplicon sequencing using the Nanopore sequencing platform for in-house sequence validation. The workflow, called DuBA.flow, is a start-to-finish procedure that provides all necessary steps from a single colony to the final easy-to-interpret sequencing report.

Nanopore strand-specific mismatch enables de novo detection of bacterial DNA modifications.

DNA modifications in bacteria present diverse types and distributions, playing crucial functional roles. Current methods for detecting bacterial DNA modifications via nanopore sequencing typically involve comparing raw current signals to a methylation-free control. In this study, we found that bacterial DNA modification induces errors in nanopore reads. And these errors are found only in one strand but not the other, showing a strand-specific bias. Leveraging this discovery, we developed Hammerhead, a pioneering pipeline designed for de novo methylation discovery that circumvents the necessity of raw signal inference and a methylation-free control. The majority (14 out of 16) of the identified motifs can be validated by raw signal comparison methods or by identifying corresponding methyltransferases in bacteria. Additionally, we included a novel polishing strategy employing duplex reads to correct modification-induced errors in bacterial genome assemblies, achieving a reduction of over 85% in such errors. In summary, Hammerhead enables users to effectively locate bacterial DNA methylation sites from nanopore FASTQ/FASTA reads, thus holds promise as a routine pipeline for a wide range of nanopore sequencing applications, such as genome assembly, metagenomic binning, decontaminating eukaryotic genome assembly, and functional analysis for DNA modifications.

Interchromosomal segmental duplication drives translocation and loss of P. falciparum histidine-rich protein 3.

Most malaria rapid diagnostic tests (RDTs) detect Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and PfHRP3, but deletions of pfhrp2 and phfrp3 genes make parasites undetectable by RDTs. We analyzed 19,313 public whole-genome-sequenced P. falciparum field samples to understand these deletions better. Pfhrp2 deletion only occurred by chromosomal breakage with subsequent telomere healing. Pfhrp3 deletions involved loss from pfhrp3 to the telomere and showed three patterns: no other associated rearrangement with evidence of telomere healing at breakpoint (Asia; Pattern 13-TARE1); associated with duplication of a chromosome 5 segment containing multidrug-resistant-1 gene (Asia; Pattern 13-5++); and most commonly, associated with duplication of a chromosome 11 segment (Americas/Africa; Pattern 13-11++). We confirmed a 13-11 hybrid chromosome with long-read sequencing, consistent with a translocation product arising from recombination between large interchromosomal ribosome-containing segmental duplications. Within most 13-11++ parasites, the duplicated chromosome 11 segments were identical. Across parasites, multiple distinct haplotype groupings were consistent with emergence due to clonal expansion of progeny from intrastrain meiotic recombination. Together, these observations suggest negative selection normally removes 13-11++pfhrp3 deletions, and specific conditions are needed for their emergence and spread including low transmission, findings that can help refine surveillance strategies.

Prokaryotic communities profiling of Indonesian hot springs using long-read Oxford Nanopore sequencing.

OBJECTIVES: Indonesia's location at the convergence of multiple tectonic plates results in a unique geomorphological feature with abundant hot springs. This study pioneers the metagenomic exploration of Indonesian hot springs, harbouring unique life forms despite high temperatures. The microbial community of hot springs is taxonomically versatile and biotechnologically valuable. 16s rRNA amplicon sequencing of the metagenome is a viable option for the microbiome investigation. This study utilized Oxford Nanopore's long-read 16 S rRNA sequencing for enhanced species identification, improved detection of rare members, and a more detailed community composition profile. DATA DESCRIPTION: Water samples were taken from three hot springs of the Bali, Indonesia (i) Angseri, 8.362503 S, 115.133452 E; (ii) Banjar, 8.210270 S, 114.967063 E; and (iii) Batur, 8.228806 S, 115.404829 E. BioLit Genomic DNA Extraction Kit (SRL, Mumbai, India) was used to isolate DNA from water samples. The quantity and quality of the DNA were determined using a NanoDrop™ spectrophotometer and a Qubit fluorometer (Thermo Fisher Scientific, USA). The library was created using Oxford Nanopore Technology kits, and the sequencing was done using Oxford Nanopore's GridION platform. All sequencing data was obtained in FASTQ files and filtered using NanoFilt software. This dataset is valuable for searching novel bacteria diversity and their existence.

Description of the complete rDNA repeat unit structure of Coturnixjaponica Temminck et Schlegel, 1849 (Aves).

Ribosomal RNA (18S, 5.8S, 28S) gene clusters in genomes form regions that consist of multiple tandem repeats. They are located on a single or several pairs of chromosomes and play an important role in the formation of the nucleolus responsible for the assembly of ribosome subunits. The rRNA gene cluster sequences are widely used for taxonomic studies, however at present, complete information on the avian rDNA repeat unit structure including intergenic spacer sequence is available only for the chicken (Gallusgallusdomesticus Linnaeus, 1758). The GC enrichment and high-order repeats peculiarities within the intergenic spacer described for the chicken rDNA cluster may be responsible for these failures. The karyotype of the Japanese quail (Coturnixjaponica Temminck et Schlegel, 1849) deserves close attention because, unlike most birds, it has three pairs of nucleolar organizer bearing chromosomes, two of which are microchromosomes enriched in repeating elements and heterochromatin that carry translocated terminal nucleolar organizers. Here we assembled and annotated the complete Japanese quail ribosomal gene cluster sequence of 21166 base pairs (GenBank under the registration tag BankIt2509210 CoturnixOK523374). This is the second deciphered avian rDNA cluster after the chicken. Despite the revealed high similarity with the chicken corresponding sequence, it has a number of specific features, which include a slightly lower degree of GC content and the presence of bendable elements in the content of both the transcribed spacer I and the non-transcribed intergenic spacer.

Nanopore sequencing with unique molecular identifiers enables accurate mutation analysis and haplotyping in the complex lipoprotein(a) KIV-2 VNTR.

BACKGROUND: Repetitive genome regions, such as variable number of tandem repeats (VNTR) or short tandem repeats (STR), are major constituents of the uncharted dark genome and evade conventional sequencing approaches. The protein-coding LPA kringle IV type-2 (KIV-2) VNTR (5.6 kb per unit, 1-40 units per allele) is a medically highly relevant example with a particularly intricate structure, multiple haplotypes, intragenic homologies, and an intra-VNTR STR. It is the primary regulator of plasma lipoprotein(a) [Lp(a)] concentrations, an important cardiovascular risk factor. Lp(a) concentrations vary widely between individuals and ancestries. Multiple variants and functional haplotypes in the LPA gene and especially in the KIV-2 VNTR strongly contribute to this variance. METHODS: We evaluated the performance of amplicon-based nanopore sequencing with unique molecular identifiers (UMI-ONT-Seq) for SNP detection, haplotype mapping, VNTR unit consensus sequence generation, and copy number estimation via coverage-corrected haplotypes quantification in the KIV-2 VNTR. We used 15 human samples and low-level mixtures (0.5 to 5%) of KIV-2 plasmids as a validation set. We then applied UMI-ONT-Seq to extract KIV-2 VNTR haplotypes in 48 multi-ancestry 1000 Genome samples and analyzed at scale a poorly characterized STR within the KIV-2 VNTR. RESULTS: UMI-ONT-Seq detected KIV-2 SNPs down to 1% variant level with high sensitivity, specificity, and precision (0.977 ± 0.018; 1.000 ± 0.0005; 0.993 ± 0.02) and accurately retrieved the full-length haplotype of each VNTR unit. Human variant levels were highly correlated with next-generation sequencing (R2 = 0.983) without bias across the whole variant level range. Six reads per UMI produced sequences of each KIV-2 unit with Q40 quality. The KIV-2 repeat number determined by coverage-corrected unique haplotype counting was in close agreement with droplet digital PCR (ddPCR), with 70% of the samples falling even within the narrow confidence interval of ddPCR. We then analyzed 62,679 intra-KIV-2 STR sequences and explored KIV-2 SNP haplotype patterns across five ancestries. CONCLUSIONS: UMI-ONT-Seq accurately retrieves the SNP haplotype and precisely quantifies the VNTR copy number of each repeat unit of the complex KIV-2 VNTR region across multiple ancestries. This study utilizes the KIV-2 VNTR, presenting a novel and potent tool for comprehensive characterization of medically relevant complex genome regions at scale.

Complete genome sequence of Serratia plymuthica SWSY-3.47.

Serratia plymuthica strain SWSY-3.47 is a Gram-negative, rod-shaped bacteria isolated for its high chitinolytic activity. Here, we report the complete genome of this strain comprised of a single circular chromosome of 5,636,345 bp with a G + C content of 56.0%.

Complete genome sequence of polylactic acid degrading Rhodopseudomonas palustris strain R1 isolated from rice field soil.

Rhodopseudomonas palustris R1 is a phototrophic, purple, non-sulfur Gram-negative bacterium first isolated from the soils of rice fields, known for its polylactic acid degradability. Here, we present the complete circular genome sequence of this bacterium, spanning 5.32 Mbp and 4,949 protein-coding sequences.

Oxford Nanopore's 2024 sequencing technology for Listeria monocytogenes outbreak detection and source attribution: progress and clone-specific challenges.

Whole genome sequencing is an essential cornerstone of pathogen surveillance and outbreak detection. Established sequencing technologies are currently being challenged by Oxford Nanopore Technologies (ONT), which offers an accessible and cost-effective alternative enabling gap-free assemblies of chromosomes and plasmids. Limited accuracy has hindered its use for investigating pathogen transmission, but recent technology updates have brought significant improvements. To evaluate its readiness for outbreak detection, we selected 78 Listeria monocytogenes isolates from diverse lineages or known epidemiological clusters for sequencing with ONT's V14 Rapid Barcoding Kit and R10.4.1 flow cells. The most accurate of several tested workflows generated assemblies with a median of one error (SNP or indel) per assembly. For 66 isolates, the cgMLST profiles from ONT-only assemblies were identical to those generated from Illumina data. Eight assemblies were of lower quality, with more than 20 erroneous sites each, primarily caused by methylations at the GAAGAC motif (5'-GAAG6mAC-3'/5'-GT4mCTTC-3'). This led to inaccurate clustering, failing to group isolates from a persistence-associated clone that carried the responsible restriction-modification system. Out of 50 methylation motifs detected among the 78 isolates, only the GAAGAC motif was linked to substantially increased error rates. Our study shows that most L. monocytogenes genomes assembled from ONT-only data are suitable for high-resolution genotyping, but further improvements of chemistries or basecallers are required for reliable routine use in outbreak and food safety investigations.

NanoCore: core-genome-based bacterial genomic surveillance and outbreak detection in healthcare facilities from Nanopore and Illumina data.

Genomic surveillance enables the early detection of pathogen transmission in healthcare facilities and contributes to the reduction of substantial patient harm. Fast turnaround times, flexible multiplexing, and low capital requirements make Nanopore sequencing well suited for genomic surveillance purposes; the analysis of Nanopore data, however, can be challenging. We present NanoCore, a user-friendly method for Nanopore-based genomic surveillance in healthcare facilities, enabling the calculation and visualization of cgMLST-like (core-genome multilocus sequence typing) sample distances directly from unassembled Nanopore reads. NanoCore implements a mapping, variant calling, and multilevel filtering strategy and also supports the analysis of Illumina data. We validated NanoCore on two 24-isolate data sets of methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecium (VRE). In the Nanopore-only mode, NanoCore-based pairwise distances between closely related isolates were near-identical to Illumina-based SeqSphere+ distances, a gold standard commercial method (average differences of 0.75 and 0.81 alleles for MRSA and VRE; sd = 0.98 and 1.00), and gave an identical clustering into closely related and non-closely related isolates. In the "hybrid" mode, in which only Nanopore data are used for some isolates and only Illumina data for others, increased average pairwise isolate distance differences were observed (average differences of 3.44 and 1.95 for MRSA and VRE, respectively; sd = 2.76 and 1.34), while clustering results remained identical. NanoCore is computationally efficient (<15 hours of wall time for the analysis of a 24-isolate data set on a workstation), available as free software, and supports installation via conda. In conclusion, NanoCore enables the effective use of the Nanopore technology for bacterial pathogen surveillance in healthcare facilities. IMPORTANCE: Genomic surveillance involves sequencing the genomes and measuring the relatedness of bacteria from different patients or locations in the same healthcare facility, enabling an improved understanding of pathogen transmission pathways and the detection of "silent" outbreaks that would otherwise go undetected. It has become an indispensable tool for the detection and prevention of healthcare-associated infections and is routinely applied by many healthcare institutions. The earlier an outbreak or transmission chain is detected, the better; in this context, the Oxford Nanopore sequencing technology has important potential advantages over traditionally used short-read sequencing technologies, because it supports "real-time" data generation and the cost-effective "on demand" sequencing of small numbers of bacterial isolates. The analysis of Nanopore sequencing data, however, can be challenging. We present NanoCore, a user-friendly software for genomic surveillance that works directly based on Nanopore sequencing reads in FASTQ format, and demonstrate that its accuracy is equivalent to traditional gold standard short read-based analyses.

Dynamic landscape of m6A modifications and related post-transcriptional events in muscle-invasive bladder cancer.

BACKGROUND: Muscle-invasive bladder carcinoma (MIBC) is a serious and more advanced stage of bladder carcinoma. N6-Methyladenosine (m6A) is a dynamic and reversible modifications that primarily affects RNA stability and alternative splicing. The dysregulation of m6A in MIBC can be potential target for clinical interventions, but there have been limited studies on m6A modifications in MIBC and their associations with post-transcriptional regulatory processes. METHODS: Paired tumor and adjacent-normal tissues were obtained from three patients with MIBC following radical cystectomy. The additional paired tissues for validation were obtained from patients underwent transurethral resection. Utilizing Nanopore direct-RNA sequencing, we characterized the m6A RNA methylation landscape in MIBC, with a focus on identifying post-transcriptional events potentially affected by changes in m6A sites. This included an examination of differential transcript usage, polyadenylation signal sites, and variations in poly(A) tail length, providing insights into the broader impact of m6A alterations on RNA processing in MIBC. RESULTS: The prognostic-related m6A genes and m6A-risk model constructed by machine learning enables the stratification of high and low-risk patients with precision. A novel m6A modification site in the 3' untranslated region (3'UTR) of IGLL5 gene were identified, characterized by a lower m6A methylation ratio, elongated poly(A) tails, and a notable bias in transcript usage. Furthermore, we discovered two particular transcripts, VWA1-203 and CEBPB-201. VWA1-203 displayed diminished m6A methylation levels, a truncated 3'UTR, and an elongated poly(A) tail, whereas CEBPB-201 showed opposite trends, highlighting the complex interplay between m6A modifications and RNA processing. Source code was provided on GitHub ( https://github.com/lelelililele/Nanopore-m6A-analysis ). CONCLUSIONS: The state-of-the-art Nanopore direct-RNA sequencing and machine learning techniques enables comprehensive identification of m6A modification and provided insights into the potential post-transcriptional regulation mechanisms on the development and progression in MIBC.

BaseNet: A transformer-based toolkit for nanopore sequencing signal decoding.

Nanopore sequencing provides a rapid, convenient and high-throughput solution for nucleic acid sequencing. Accurate basecalling in nanopore sequencing is crucial for downstream analysis. Traditional approaches such as Hidden Markov Models (HMM), Recurrent Neural Networks (RNN), and Convolutional Neural Networks (CNN) have improved basecalling accuracy but there is a continuous need for higher accuracy and reliability. In this study, we introduce BaseNet (https://github.com/liqingwen98/BaseNet), an open-source toolkit that utilizes transformer models for advanced signal decoding in nanopore sequencing. BaseNet incorporates both autoregressive and non-autoregressive transformer-based decoding mechanisms, offering state-of-the-art algorithms freely accessible for future improvement. Our research indicates that cross-attention weights effectively map the relationship between current signals and base sequences, joint loss training through adding a pair of forward and reverse decoder facilitate model converge, and large-scale pre-trained models achieve superior decoding accuracy. This study helps to advance the field of nanopore sequencing signal decoding, contributes to technological advancements, and provides novel concepts and tools for researchers and practitioners.

Characterizing a lethal CAG-ACE2 transgenic mouse model for SARS-CoV-2 infection using Cas9-enhanced nanopore sequencing.

The SARS-CoV-2 pandemic has underscored the necessity for functional transgenic animal models for testing. Mouse lines with overexpression of the human receptor ACE2 serve as the common animal model to study COVID-19 infection. Overexpression of ACE2 under a strong ubiquitous promoter facilitates convenient and sensitive testing of COVID-19 pathology. We performed pronuclear microinjections using a 5 kb CAG-ACE2 linear transgene construct and identified three founder lines with 140, 72, and 73 copies, respectively. Two of these lines were further analyzed for ACE2 expression profiles and sensitivity to SARS-CoV-2 infection. Both lines expressed ACE2 in all organs analyzed. Embryonic fibroblast cell lines derived from transgenic embryos demonstrated severe cytopathic effects following infection, even at low doses of SARS-CoV-2 (0,1-1.0 TCID50). Infected mice from the two lines began to show COVID-19 clinical signs three days post-infection and succumbed between days 4 and 7. Histological examination of lung tissues from terminally ill mice revealed severe pathological alterations. To further characterize the integration site in one of the lines, we applied nanopore sequencing combined with Cas9 enrichment to examine the internal transgene concatemer structure. Oxford Nanopore sequencing (ONT) is becoming the gold standard for transgene insert characterization, but it is relatively inefficient without targeted region enrichment. We digested genomic DNA with Cas9 and gRNA against the ACE2 transgene to create ends suitable for ONT adapter ligation. ONT data analysis revealed that most of the transgene copies were arranged in a head-to-tail configuration, with palindromic junctions being rare. We also detected occasional plasmid backbone fragments within the concatemer, likely co-purified during transgene gel extraction, which is a common occurrence in pronuclear microinjections.

RAPiD: a rapid and accurate plant pathogen identification pipeline for on-site nanopore sequencing.

Nanopore sequencing technology has enabled the rapid, on-site taxonomic identification of samples from anything and anywhere. However, sequencing errors, inadequate databases, as well as the need for bioinformatic expertise and powerful computing resources, have hampered the widespread use of the technology for pathogen identification in the agricultural sector. Here we present RAPiD, a lightweight and accurate real-time taxonomic profiling pipeline. Compared to other metagenomic profilers, RAPiD had a higher classification precision achieved through the use of a curated, non-redundant database of common agricultural pathogens and extensive quality filtering of alignments. On a fungal, bacterial and mixed mock community RAPiD was the only pipeline to detect all members of the communities. We also present a protocol for in-field sample processing enabling pathogen identification from plant sample to sequence within 3 h using low-cost equipment. With sequencing costs continuing to decrease and more high-quality reference genomes becoming available, nanopore sequencing provides a viable method for rapid and accurate pathogen identification in the field. A web implementation of the RAPiD pipeline for real-time analysis is available at https://agrifuture.senckenberg.de.

Evaluation of Commercial RNA Extraction Protocols for Avian Influenza Virus Using Nanopore Metagenomic Sequencing.

Avian influenza virus (AIV) is a significant threat to the poultry industry, necessitating rapid and accurate diagnosis. The current AIV diagnostic process relies on virus identification via real-time reverse transcription-polymerase chain reaction (rRT-PCR). Subsequently, the virus is further characterized using genome sequencing. This two-step diagnostic process takes days to weeks, but it can be expedited by using novel sequencing technologies. We aim to optimize and validate nucleic acid extraction as the first step to establishing Oxford Nanopore Technologies (ONT) as a rapid diagnostic tool for identifying and characterizing AIV from clinical samples. This study compared four commercially available RNA extraction protocols using AIV-known-positive clinical samples. The extracted RNA was evaluated using total RNA concentration, viral copies as measured by rRT-PCR, and purity as measured by a 260/280 absorbance ratio. After NGS testing, the number of total and influenza-specific reads and quality scores of the generated sequences were assessed. The results showed that no protocol outperformed the others on all parameters measured; however, the magnetic particle-based method was the most consistent regarding CT value, purity, total yield, and AIV reads, and it was less error-prone. This study highlights how different RNA extraction protocols influence ONT sequencing performance.

Multiplex PCR Approach for Rapid African Swine Fever Virus Genotyping.

African swine fever virus (ASFV) has been spreading through Europe, Asia, and the Caribbean after its introduction in Georgia in 2007 and, due to its particularly high mortality rate, poses a continuous threat to the pig industry. The golden standard to trace back the ASFV is whole genome sequencing, but it is a cost and time-intensive methodology. A more efficient way of tracing the virus is to amplify only specific genomic regions relevant for genotyping. This is mainly accomplished by amplifying single amplicons by PCR followed by Sanger sequencing. To reduce costs and processivity time, we evaluated a multiplex PCR based on the four primer sets routinely used for ASFV genotyping (B646L, E183L, B602L, and intergenic I73R-I329L), which was followed by Nanopore ligation-based amplicon sequencing. We show that with this protocol, we can genotype ASFV DNA originating from different biological matrices and correctly classify multiple genotypes and strains using a single PCR reaction. Further optimization of this method can be accomplished by adding or swapping the primer sets used for amplification based on the needs of a specific country or region, making it a versatile tool that can speed up the processing time and lower the costs of genotyping during ASFV outbreaks.

Combining Short- and Long-Read Sequencing Technologies to Identify SARS-CoV-2 Variants in Wastewater.

During the COVID-19 pandemic, the monitoring of SARS-CoV-2 RNA in wastewater was used to track the evolution and emergence of variant lineages and gauge infection levels in the community, informing appropriate public health responses without relying solely on clinical testing. As more sublineages were discovered, it increased the difficulty in identifying distinct variants in a mixed population sample, particularly those without a known lineage. Here, we compare the sequencing technology from Illumina and from Oxford Nanopore Technologies, in order to determine their efficacy at detecting variants of differing abundance, using 248 wastewater samples from various Quebec and Ontario cities. Our study used two analytical approaches to identify the main variants in the samples: the presence of signature and marker mutations and the co-occurrence of signature mutations within the same amplicon. We observed that each sequencing method detected certain variants at different frequencies as each method preferentially detects mutations of distinct variants. Illumina sequencing detected more mutations with a predominant lineage that is in low abundance across the population or unknown for that time period, while Nanopore sequencing had a higher detection rate of mutations that are predominantly found in the high abundance B.1.1.7 (Alpha) lineage as well as a higher sequencing rate of co-occurring mutations in the same amplicon. We present a workflow that integrates short-read and long-read sequencing to improve the detection of SARS-CoV-2 variant lineages in mixed population samples, such as wastewater.

Complete genome sequence of Mediterraneibacter gnavus strain RI1, isolated from human feces.

We report the complete genome sequence of Mediterraneibacter gnavus strain RI1, a Gram-positive anaerobic gut microbe isolated from human feces. The complete circular genome has a genome size of 3.25 Mb, with a G+C content of 42.6%.

New insights into the dynamics of m6A epitranscriptome: hybrid-seq identifies novel mRNAs of the m6A writers METTL3/14.

Background: N6-methyladenosine (m6A), a prevalent mRNA modification, is dynamically regulated by methyltransferases, including METTL3 and METTL14.Materials & methods: In the current study, we employed a custom hybrid-seq method to identify novel METTL3/14 transcripts, explore their protein-coding capacities and predict the putative role of the METTL isoforms.Results: Demultiplexing of the hybrid-seq barcoded datasets unraveled the expression patterns of the newly identified mRNAs in major malignancies as well as in non-malignant cells, providing a deeper understanding of the methylation pathways. Open reading frame query revealed novel METTL3/14 isoforms, broadening our perspective for the structural diversity within METTL family.Conclusion: Our findings offer significant insights into the intricate transcriptional landscape of METTL3/14, shedding light on the regulatory mechanisms underlying methylation in mRNAs.

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Draft genome sequence of Taiwanofungus gaoligongensis strain YAF008 isolated from Gaoligong Mountain.

The draft genome sequence of Taiwanofungus gaoligongensis YAF008 was reported. The genome size of T. gaoligongensis YAF008 was 34.7M bp with 50.72% GC content. The genome resource will support future research into potential secondary metabolite diversity of this fungus.