Journal: Nature Communications

Article Title: Diverse monogenic subforms of human spermatogenic failure

doi: 10.1038/s41467-022-35661-z

Figure Lengend Snippet: a A schematic of piRNA biogenesis with the components affected among NOA cases in this study highlighted in red. piRNAs are produced by two biogenesis pathways. The primary pathway involves transcription of long precursor-transcripts from genomic clusters, which are then processed in the cytoplasm. b Cell scores for component 59, which encodes both piRNA processing genes and target pre-pre-piRNAs, indicate that it is expressed primarily in pre-leptotene spermatocytes. c Gene loadings for component 59. Note TDRD10 (black box), a protein-coding gene that has yet to be characterized for a role in mammalian piRNA processing. d H&E stain of TDRD9 patient biopsy showing spermatogenic arrest. The most mature germ cell observed were early round spermatids, which often appeared multinucleated (arrows). In addition, many pyknotic cells were observed (arrowheads). Scale bar = 50 µm. e Size distribution of piRNAs detected in the NOA case with a biallelic missense variant in TDRD12 and matching control, derived from small RNA-sequencing of testis tissue. f Significantly decreased fraction of mature (<32 bases) piRNAs was detected in testis of patients with piRNA pathway mutations, indicative of faulty processing of the immature piRNA transcripts.

Article Snippet: Small RNA sequencing libraries were prepared using CATS Small RNA-seq kit (Diagenode, Cat. #C05010040) and sequenced on the MiSeq platform (Illumina, San Diego, CA).

Techniques: Produced, Staining, Variant Assay, Derivative Assay, RNA Sequencing Assay

Journal: Journal of Biomolecular Techniques : JBT

Article Title: Multisite Evaluation of Next-Generation Methods for Small RNA Quantification

doi: 10.7171/jbt.20-3102-001

Figure Lengend Snippet: Study design and workflow metrics. A) Schematic of study design is shown. MUR, MUR-D, and HBR were processed using 9 different smRNA profiling methods at 4 sites each. The general methodologies included sequential ligation (Illumina, TriLink, Qiagen, NEB, PerkinElmer, Lexogen), template switching (Takara, Diagenode), circularization (Somagenics), and NanoString probe-based hybridization. B) Total start-to-finish preparation time for each kit as reported by sites. C) Mean “ease of use” reported by each site (scale: 1 = uncomfortable, 5 = comfortable). D) Library preparation success rates. Light color blocks are successfully produced libraries. Dark colors are failed libraries. CLO and CLO-S, Takara Bio (Clontech) SMARTer smRNA-Seq Kit; DIA, Diagenode CATS Small RNA-Seq Kit; ILL and ILMN, Illumina TruSeq Small RNA Library Prep Kit; LEX, Lexogen Small RNA-Seq Library Prep Kit; NANO, NanoString nCounter miRNA Expression Assay; NEB, New England Biolabs NEBNext Small RNA Library Prep Set; PEB, PerkinElmer NextFlex Small RNA-Seq Kit v.3; QIA, Qiagen QIAseq miRNA Library Kit; SOM, Somagenics RealSeq-AC miRNA Library Kit; TRI, Trilink CleanTag Small RNA Library Prep Kit.

Article Snippet: Diagenode CATS Small RNA-Seq Kit (kit C05010044, protocol v.2|09.17; Diagenode, Liège, Belgium) The amount of input RNA for HBR and MUR-D was 10 ng and 35 pg for MUR.

Techniques: Ligation, Hybridization, Produced, RNA Sequencing Assay, Expressing

Journal: Journal of Biomolecular Techniques : JBT

Article Title: Multisite Evaluation of Next-Generation Methods for Small RNA Quantification

doi: 10.7171/jbt.20-3102-001

Figure Lengend Snippet: miRNA detection and bias. Percentages of Miltenyi Biotec miRXplore miRNA detected above 5 CPM in MUR (A) and in MUR-D (B) are shown. Percentages and amplitude of Miltenyi Biotec miRXplore miRNA detected that deviated from the median are shown in MUR (C) or MUR-D (D). The darkest shade is within 2-fold of the median; the medium shade is 2–10-fold either up or down vs. the expected value; the lightest shade is >10× either increased or decreased. Percentages of reads increased >10× from median in MUR (E) or MUR-D (F). CLO-S and C, Takara Bio (Clontech) SMARTer smRNA-Seq Kit; DIA and D, Diagenode CATS Small RNA-Seq Kit; ILMN and I, Illumina TruSeq Small RNA Library Prep Kit; LEX and L, Lexogen Small RNA-Seq Library Prep Kit; Nano, NanoString nCounter miRNA Expression Assay; NEB and N, New England Biolabs NEBNext Small RNA Library Prep Set; PEB and P, PerkinElmer NextFlex Small RNA-Seq Kit v.3; QIA and Q, Qiagen QIAseq miRNA Library Kit; SOM and S, Somagenics RealSeq-AC miRNA Library Kit; TRI and T, Trilink CleanTag Small RNA Library Prep Kit.

Article Snippet: Diagenode CATS Small RNA-Seq Kit (kit C05010044, protocol v.2|09.17; Diagenode, Liège, Belgium) The amount of input RNA for HBR and MUR-D was 10 ng and 35 pg for MUR.

Techniques: RNA Sequencing Assay, Expressing

Journal: Journal of Biomolecular Techniques : JBT

Article Title: Multisite Evaluation of Next-Generation Methods for Small RNA Quantification

doi: 10.7171/jbt.20-3102-001

Figure Lengend Snippet: Relative expression of miRNAs. A) Correlation scatter plots of miRNAs detected in MUR and MUR-D for the QIA method, with miRNAs scored in log2 mapped reads per million. B) Correlation scatter plot of miRNAs detected in MUR-D using LEX vs. ILL kits, with miRNAs scored in log2 mapped reads per million. C) Unsupervised hierarchical clustering heatmap of log2 transformed CPM across all methods. C (prep kit), Takara Bio (Clontech) SMARTer smRNA-Seq Kit; C (prep type), circularization; D, Diagenode CATS Small RNA-Seq Kit; ILL and I, Illumina TruSeq Small RNA Library Prep Kit; L (prep type), sequential ligation; LEX and L (prep kit), Lexogen Small RNA-Seq Library Prep Kit; N, New England Biolabs NEBNext Small RNA Library Prep Set; Nano, NanoString nCounter miRNA Expression Assay; NO, No size selection; P, PerkinElmer NextFlex Small RNA-Seq Kit v.3; QIA and Q, Qiagen QIAseq miRNA Library Kit; S, Somagenics RealSeq-AC miRNA Library Kit; SAGE, pippin prep; SPRI, solid phase reversible immobilization; T (prep kit), Trilink CleanTag Small Library Prep Kit; T (prep type), template switching.

Article Snippet: Diagenode CATS Small RNA-Seq Kit (kit C05010044, protocol v.2|09.17; Diagenode, Liège, Belgium) The amount of input RNA for HBR and MUR-D was 10 ng and 35 pg for MUR.

Techniques: Expressing, Transformation Assay, RNA Sequencing Assay, Ligation, Selection

Journal: Journal of Biomolecular Techniques : JBT

Article Title: Multisite Evaluation of Next-Generation Methods for Small RNA Quantification

doi: 10.7171/jbt.20-3102-001

Figure Lengend Snippet: Detection of smRNAs in complex samples. Insert sizes in MUR-D (A) and HBR (B) are shown with inserts <18 nt as white, 18–23 nt as the lightest shade, 24–35 nt as the medium shade, and >35 nt as the darkest shade. Data from the different method types are colored (ligation, blue; polyA tailing, orange; circularization, green). The right panels show the smRNA target distribution for MUR-D (C) and HBR (D). Mapped targets are noted in the key. Unmapped targets are shown in black. C, Takara Bio (Clontech) SMARTer smRNA-Seq Kit; D, Diagenode CATS Small RNA-Seq Kit; I, Illumina TruSeq Small RNA Library Prep Kit; L, Lexogen Small RNA-Seq Library Prep Kit; lincRNA, long intervening noncoding RNA; N, New England Biolabs NEBNext Small RNA Library Prep Set; ncRNA, noncoding RNA; P, PerkinElmer NextFlex Small RNA-Seq Kit v.3; Q, Qiagen QIAseq miRNA Library Kit; S, Somagenics RealSeq-AC miRNA Library Kit; snoRNA, small nucleolar RNA; snRNA, small nuclear RNA; T, Trilink CleanTag Small Library Prep Kit.

Article Snippet: Diagenode CATS Small RNA-Seq Kit (kit C05010044, protocol v.2|09.17; Diagenode, Liège, Belgium) The amount of input RNA for HBR and MUR-D was 10 ng and 35 pg for MUR.

Techniques: Ligation, RNA Sequencing Assay