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Method: Analysis of coding gene expression from small RNA sequencing
- Aygun Azadova,
- Anthonia Ekperuoh,
- Greg N. Brooke,
- and Antonio Marco
Genome Res. March 2026 36: 611-618; Published in Advance February 10, 2026, doi:10.1101/gr.281364.125
...genomics. Early works in molecular biology on gene expression were limited, because purified RNA is unstable and difficult to work with. However, the discovery (and use in the laboratory) of reverse transcriptase, permitting the controlled synthesis of RNAs into cDNAs (Maniatis et al. 1976...
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Research: Dynamics and consequences of differential RNA isoform production during cardiomyocyte fate determination and early-stage maturation
- Joanna Delimata-Raczek,
- Natalia Koralewska,
- Bart Krist,
- Magdalena Rakoczy,
- Marek Figlerowicz,
- and Ireneusz Stolarek
Genome Res. April 3, 2026 ; Published in Advance April 3, 2026, doi:10.1101/gr.281150.125
...of differentiation efficiency. All cells in culture were maintained at 37°C in incubators with 5% CO2. RNA extraction and cDNA synthesis The RNA was extracted from the iCMs on day 4 and day 30. Cells were first washedwith PBS and dissociated from the plate either by 3 min incubation in 37°C with PBS with EDTA (4 day...
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Research: Dynamic A-to-I RNA editing in response to gut microbiome in honeybees
- Yuange Duan,
- Min Huang,
- Lin Ye,
- Ling Ma,
- Yashuai Wu,
- Yating Du,
- Jiyao Liu,
- Fan Song,
- Li Tian,
- Wanzhi Cai,
- Hu Li,
- Xin Zhou,
- and Shiqi Luo
Genome Res. March 4, 2026 ; Published in Advance March 4, 2026, doi:10.1101/gr.280291.124
...sites by Sanger sequencing For the p.482 Ile >Met editing site in Adar, we conducted amplification ofAdar from both gDNA and cDNA extracted from the same sample (the first batch data of three CV and three GF). For cDNA synthesis, 1 µg of total RNA was incubated with gDNA wiper (Vazyme Biotech) for 2 min...
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Method: Deep structural clustering reveals hidden systematic biases in RNA sequencing data
- Qiang Su,
- Yi Long,
- Deming Gou,
- Junmin Quan,
- Xiaoming Zhou,
- and Qizhou Lian
Genome Res. November 2025 35: 2563-2577; Published in Advance September 19, 2025, doi:10.1101/gr.280713.125
...regions, influenced by the unique tertiary structures of these regions, thereby affecting the sequencing output. In contrast, linear regression analysis was not applicable to data obtained from the oligo(dT)-based protocol. This method uses a poly(T) primer to initiate cDNA synthesis, focusing almost...
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Method: Tissular chromatin-state cartography based on double-barcoded DNA arrays that capture unloaded PA-Tn5 transposase
- Maria Grazia Mendoza-Ferri,
- Gwendoline Lozachmeur,
- Maximilien Duvina,
- Laetitia Perret,
- Didier Merciris,
- Anne Gigout,
- and Marco Antonio Mendoza-Parra
Genome Res. July 2025 35: 1633-1645; Published in Advance May 13, 2025, doi:10.1101/gr.280305.124
...by magnesium chloride, leading to the formation of hairpin-like DNA structures. (E) Scheme illustrating the molecular biology steps required for generating a copy of the genomic DNA captured by the DNA probes, followed by their amplification for Illumina massive parallel DNA sequencing.Printed DNA arrays...
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Method: Rapid and accurate demultiplexing of direct RNA nanopore sequencing data with SeqTagger
- Leszek P. Pryszcz,
- Gregor Diensthuber,
- Laia Llovera,
- Rebeca Medina,
- Anna Delgado-Tejedor,
- Luca Cozzuto,
- Julia Ponomarenko,
- and Eva Maria Novoa
Genome Res. April 2025 35: 956-966; Published in Advance January 29, 2025, doi:10.1101/gr.279290.124
...for polymerase chain reaction (PCR) amplification and cDNA synthesis steps, thus preserving the native state of RNA molecules and providing a less biased and comprehensive view of the transcriptome (Workman et al. 2020).However, key challenges persist, such as the lack of commercial barcoding kits...
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Method: Tn5 tagments and transposes oligos to single-stranded DNA for strand-specific RNA sequencing
- Yanjun Zhang,
- Yin Tang,
- Zhongxing Sun,
- Junqi Jia,
- Yuan Fang,
- Xinyi Wan,
- and Dong Fang
Genome Res. March 2023 33: 412-426; Published in Advance March 23, 2023, doi:10.1101/gr.277213.122
...to ssDNA, we proposed that Tn5 was capable of constructing the strand-specific RNA sequencing library from single-strand cDNA, which omitted the second-strand synthesis.Many different high-throughput RNA sequencing technologies have been developed to profile enriched RNA (Stark et al. 2019). Fragmented...
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Resource: Full-length RNA transcript sequencing traces brain isoform diversity in house mouse natural populations
- Wenyu Zhang,
- Anja Guenther,
- Yuanxiao Gao,
- Kristian Ullrich,
- Bruno Huettel,
- Aftab Ahmad,
- Lei Duan,
- Kaizong Wei,
- and Diethard Tautz
Genome Res. November 2024 34: 2118-2132; Published in Advance September 17, 2024, doi:10.1101/gr.279166.124
...with hallmarks of both, a 5′ CAP and a poly(A) tail. It has been reported that the TeloPrime Full-Length cDNA Amplification Kit could selectively synthesize cDNA molecules from mRNAs carrying a 5′ CAP, in complement to the standard PacBio cDNA library preparation protocol that could only selectively synthesize cDNA...
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Method: Native molecule sequencing by nano-ID reveals synthesis and stability of RNA isoforms
- Kerstin C. Maier,
- Saskia Gressel,
- Patrick Cramer,
- and Björn Schwalb
Genome Res. September 2020 30: 1332-1344; Published in Advance September 4, 2020, doi:10.1101/gr.257857.119
...(Shendure et al. 2017). Finally, labeled RNA purification (TT-seq) and cDNA library preparation (TT-seq, SLAM seq, and TimeLapse-seq) can also introduce biases, such as cross-contamination with unlabeled RNA and read duplication.Therefore, monitoring RNA metabolism at the level of RNA isoforms requires...
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Research: Cytosolic and mitochondrial translation elongation are coordinated through the molecular chaperone TRAP1 for the synthesis and import of mitochondrial proteins
- Rosario Avolio,
- Ilenia Agliarulo,
- Daniela Criscuolo,
- Daniela Sarnataro,
- Margherita Auriemma,
- Sabrina De Lella,
- Sara Pennacchio,
- Giovanni Calice,
- Martin Y. Ng,
- Carlotta Giorgi,
- Paolo Pinton,
- Barry S. Cooperman,
- Matteo Landriscina,
- Franca Esposito,
- and Danilo Swann Matassa
Genome Res. August 2023 33: 1242-1257; Published in Advance July 24, 2023, doi:10.1101/gr.277755.123
...mitochondrial and cytosolic translation, as well as in mitochondrial respiration.The flow of genetic information from DNA to protein entails multiple highly regulated steps from mRNA transcription, processing, and export to translation into proteins with subsequent folding, post-translational modification, and...
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