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Pre-miRNA

Fig. 2 RNAi inducers used in antiviral strategies. In general, RNAi is induced either by transfection of synthetic siRNAs into cells, or by stable or transient intracellular expression of double-stranded siRNA precursors (shRNA, e-shRNA, IhRNA, or pri-miRNAs). After transcription in the nucleus shRNAs, IhRNAs and e-shRNAs are exported to the cytoplasm and subsequently diced into mature siRNAs. Pri-miRNAs modified to encode antiviral siRNAs first undergo cleavage by Drosha before they are exported to the cytoplasm. Here the antiviral pre-miRNAs (also called shRNA-miRs) are processed by Dicer into the mature miRNAs. After loading of the antisense strand of the siRNAs/miRNAs into RISC, the complex will target and cleave viral transcripts bearing the complementary sequences... Fig. 2 RNAi inducers used in antiviral strategies. In general, RNAi is induced either by transfection of synthetic siRNAs into cells, or by stable or transient intracellular expression of double-stranded siRNA precursors (shRNA, e-shRNA, IhRNA, or pri-miRNAs). After transcription in the nucleus shRNAs, IhRNAs and e-shRNAs are exported to the cytoplasm and subsequently diced into mature siRNAs. Pri-miRNAs modified to encode antiviral siRNAs first undergo cleavage by Drosha before they are exported to the cytoplasm. Here the antiviral pre-miRNAs (also called shRNA-miRs) are processed by Dicer into the mature miRNAs. After loading of the antisense strand of the siRNAs/miRNAs into RISC, the complex will target and cleave viral transcripts bearing the complementary sequences...
Quantitation of the mature miRNA in its functionally active form is often performed rather than that of the pri-miRNA or pre-miRNA. However, it is sometimes of interest to investigate pri-miRNA expression in order to elucidate transcriptional control of miRNA expression. In addition, combined differential expression analysis of pri-miRNAs, pre-miRNAs, and mature miRNAs may determine whether altered mature miRNA expression is a result of regulation in miRNA biogenesis. Many commercially available assays for qPCR analysis of pri- and pre-miRNAs are available. The first step in qPCR analysis of miRNAs involves the RT of RNA to generate a cDNA sequence. Because of the small size of mature miRNAs and their lack of a poly(A) tail,... [Pg.30]

Fig. 3 The microRNA (miRNA) pathway. An miRNA is first transcribed as part of an imperfect hairpin in a longer Pol II transcript with 5 cap (open circle) and polyadenosine tail (AAAAAA). The hairpin (pre-miRNA) is removed from the transcript by the nuclear RNase III type enzyme Drosha and its partner, DGCR8. Exportin-5 (Xpo-5) transports the resulting by-product to the cytoplasm, where Dicer liberates a short-lived miRNA duplex. The duplex is unwound, and one strand enters RISC. If the miRNA is mismatched to its target (left), it does not induce cleavage, but may inhibit translation. With a perfect or near-perfect match (right), the target RNA is destroyed by RISC. Fig. 3 The microRNA (miRNA) pathway. An miRNA is first transcribed as part of an imperfect hairpin in a longer Pol II transcript with 5 cap (open circle) and polyadenosine tail (AAAAAA). The hairpin (pre-miRNA) is removed from the transcript by the nuclear RNase III type enzyme Drosha and its partner, DGCR8. Exportin-5 (Xpo-5) transports the resulting by-product to the cytoplasm, where Dicer liberates a short-lived miRNA duplex. The duplex is unwound, and one strand enters RISC. If the miRNA is mismatched to its target (left), it does not induce cleavage, but may inhibit translation. With a perfect or near-perfect match (right), the target RNA is destroyed by RISC.
The therapeutic effects mediated by exosomes from human stem cells are numerous. Exosomes from human CD34(-I-) stem cells mediate proangiogenic paracrine activity to help rebuild vasculature (Sahoo et al., 2011). Adult stem-cell-derived exosomes also shuttle selected patterns of mRNA, miRNA, and pre-miRNA associated with several cellular functions involved in the control of transcription, proliferation, and cell immune regulation. [Pg.199]

Shi W, Hendrix D, Levine M, Haley B. A distinct class of small RNAs arises from pre-miRNA-proximal region in a simple cordate. Nat Struct Mol Biol. 2009 16 183-9. [Pg.658]

To assess the biological activity of the chemically synthesized 110-nt pre-miRNA candidate, we measured its specific gene-silencing effect by two methods, mRNA-... [Pg.29]

Fig. 8 Pre-miRNA candidate pre-miR-196a-2. The grey part indicates the sequence of miRNA-196-a [111]... Fig. 8 Pre-miRNA candidate pre-miR-196a-2. The grey part indicates the sequence of miRNA-196-a [111]...
Fig. 11 HOXC8 mRNA expression analysis by RT-PCR PCS cells expressing HOXC8 were transfected with the 110-nt pre-miRNA or the 22-nt iniRNA duplex and HOXC8 mRNA was determined by RT-PCR after 24 h. HOXC8 mRNA expression was normalized to that of glyceral-dehyde-3-phosphate dehydrogenase [117]... Fig. 11 HOXC8 mRNA expression analysis by RT-PCR PCS cells expressing HOXC8 were transfected with the 110-nt pre-miRNA or the 22-nt iniRNA duplex and HOXC8 mRNA was determined by RT-PCR after 24 h. HOXC8 mRNA expression was normalized to that of glyceral-dehyde-3-phosphate dehydrogenase [117]...
Fig. 12 Gene-silencing effect of chemically synthesized pre-nuRNA. Left-. Effect of pre-nuRNA on expression of luciferase target gene. G3T-hi cells were transfected with pHOXB-Luc reporter plasmid, and effector RNA (100 nM) and reporter assays performed 48 h after transfection. Each value shown is the average of three independent experiments, and standard deviations are indicated as error bars. Right. Sense/antisense suppression activity ratios. G3T-hi cells were transfected with pHOXB-Luc or pHOXB-Luc-antisense reporter plasmid and effector RNA (30 nM), and reporter assays performed 48 h after transfection. The results are presented as the ratio of the average percent suppression of luciferase-containing sense and antisense target by the 110-nt pre-miRNA or the 22-nt miRNA duplex [111]... Fig. 12 Gene-silencing effect of chemically synthesized pre-nuRNA. Left-. Effect of pre-nuRNA on expression of luciferase target gene. G3T-hi cells were transfected with pHOXB-Luc reporter plasmid, and effector RNA (100 nM) and reporter assays performed 48 h after transfection. Each value shown is the average of three independent experiments, and standard deviations are indicated as error bars. Right. Sense/antisense suppression activity ratios. G3T-hi cells were transfected with pHOXB-Luc or pHOXB-Luc-antisense reporter plasmid and effector RNA (30 nM), and reporter assays performed 48 h after transfection. The results are presented as the ratio of the average percent suppression of luciferase-containing sense and antisense target by the 110-nt pre-miRNA or the 22-nt miRNA duplex [111]...
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MiRNAs

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