• Title/Summary/Keyword: G-rich RNA

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Secondary Structure for RNA Aptamers Binding to Guanine-Rich Sequence in the 5'-UTR RNA of N-Ras Oncogene

  • Cho, Bongrae
    • Journal of the Korean Chemical Society
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    • v.65 no.2
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    • pp.121-124
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    • 2021
  • RNA molecules which bind to the G-rich sequence in the 5'-UTR RNA which plays an important role in expression of N-ras, were selected. The secondary structures of five selected RNA aptamers including primer sequence were found by the CLC RNA workbench ver. 4.2 program (www.clcbio.com) and investigated with RNA structural probes such as RNase T1 which has specificity for a G in single-stranded region, RNase V1 specific for double strand and nuclease S1 specific for single strand. The generalized secondary structure model was proposed and characterized. It was composed of a central long double strand region flanked by single strand region at both end sides. The double strand region had an internal single-strand region and bulges. The single strand loop in the right side was composed of four or five nucleotides.

An Improved Total RNA Extraction Method for White Jelly Mushroom Tremella fuciformis Rich in Polysaccharides

  • Zhu, Hanyu;Sun, Xueyan;Liu, Dongmei;Zheng, Liesheng;Chen, Liguo;Ma, Aimin
    • Mycobiology
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    • v.45 no.4
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    • pp.434-437
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    • 2017
  • An improved method for extracting high quality and quantity RNA from a jelly mushroom and a dimorphic fungus-Tremella fuciformis which is especially rich in polysaccharides, is described. RNA was extracted from T. fuciformis mycelium M1332 and its parental monokaryotic yeast-like cells Y13 and Y32. The A260/280 and A260/230 ratios were both approximately 2, and the RNA integrity number was larger than 8.9. The yields of RNA were between 108 and $213{\mu}g/g$ fresh wt. Downstream molecular applications including reverse transcriptional PCR and quantitative real-time PCR were also performed. This protocol is reliable and may be widely applicable for total RNA extraction from other jelly mushrooms or filamentous fungi rich in polysaccharides.

Flooding Stress-Induced Glycine-Rich RNA-Binding Protein from Nicotiana tabacum

  • Lee, Mi-Ok;Kim, Keun Pill;Kim, Byung-gee;Hahn, Ji-Sook;Hong, Choo Bong
    • Molecules and Cells
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    • v.27 no.1
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    • pp.47-54
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    • 2009
  • A cDNA clone for a transcript preferentially expressed during an early phase of flooding was isolated from Nicotiana tabacum. Nucleotide sequencing of the cDNA clone identified an open reading frame that has high homology to the previously reported glycine-rich RNA-binding proteins. The open reading frame consists of 157 amino acids with an N-terminal RNA-recognition motif and a C-terminal glycine-rich domain, and thus the cDNA clone was designated as Nicotiana tabaccum glycine-rich RNA-binding protein-1 (NtGRP1). Expression of NtGRP1 was upregulated under flooding stress and also increased, but at much lower levels, under conditions of cold, drought, heat, high salt content, and abscisic acid treatment. RNA homopolymer-binding assay showed that NtGRP1 binds to all the RNA homopolymers tested with a higher affinity to poly r(G) and poly r(A) than to poly r(U) and poly r(C). Nucleic acid-binding assays showed that NtGRP1 binds to ssDNA, dsDNA, and mRNA. NtGRP1 suppressed expression of the fire luciferase gene in vitro, and the suppression of luciferase gene expression could be rescued by addition of oligonucleotides. Collectively, the data suggest NtGRP1 as a negative modulator of gene expression by binding to DNA or RNA in bulk that could be advantageous for plants in a stress condition like flooding.

Methionyl-tRNA-synthetase is a Novel Interacting Protein of LRRK2 (파킨스병 유전인자인 LRRK2와 상호작용하는 methionyl-tRNA synthetase)

  • Kim, Hyejung;Ho, Dong Hwan;Son, Ilhong;Seol, Wongi
    • Journal of Life Science
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    • v.28 no.2
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    • pp.170-175
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    • 2018
  • Parkinson's disease (PD) is the most common movement disorder and the second most common neurodegenerative disease after Alzheimer's disease. Approximately 5~10% of PD patients are familial PD cases. Leucine-rich repeat kinase 2 (LRRK2) has been known to be a causal gene of PD when it is mutated. LRRK2 contains the functional kinase and GTPase domains as well as leucine-rich repeat (LRR) and WD40 domains that are known to play critical roles for protein-protein interaction, suggesting that LRRK2-interacting proteins are important regulators for PD pathogenesis. In an effort to identify proteins interacting with LRRK2, we carried out co-immunoprecipitation of LRRK2 antibody using extracts of NIH3T3 cells that express LRRK2 at a relatively high level. The mass spectrometry analysis of a precipitated band revealed that the co-precipitated band was methionyl-tRNA synthetase (MRS), an ancient enzyme that transfers methionin to its cognate tRNA. The interaction of MRS with LRRK2 was confirmed again by co-immunoprecipitation of endogenous proteins and GST pull-down assays. However, LRRK2 did not phosphorylate recombinant MRS protein in in vitro kinase assays, and over-expression of LRRK2 or MRS did not affect the stability of its partner protein. Our data indicate that LRRK2 interacts with but does not phosphorylate MRS, and the stability of each partner is not affected by the other.

Effects of the cis-Acting Element in the 3' End of Porcine $\beta$-Casein Gene on the Expression in Mammary Epithelial Cells (돼지 $\beta$-Casein 유전자의 3' 말단 부위의 cis-Acting Element가 유선 상피 세포내의 발현에 미치는 영향)

  • Lee, Hwi-Cheul;Kim, Byoung-Ju;Byun, Sung-June;Lee, Seung-Hoon;Kim, Min-Ji;Chung, Hee Kyoung;Lee, Hyun-Gi;Jo, Su-Jin;Chang, Won-Kyong;Park, Jin-Ki;Lee, Poong-Yeon
    • Reproductive and Developmental Biology
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    • v.32 no.3
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    • pp.153-158
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    • 2008
  • Tissue-specific and temporal regulation of milk protein gene expression is advantageous when creating transgenic animal that produces foreign protein into milk. Gene expression, i.e. protein production, is regulated not only by promoter strength but also mRNA stability. Especially, poly A tail length by polyadenylation affects in vivo and in vitro mRNA stability and translation efficiency of the target gene. In the present study, nucleotide sequence of 3'-UTR was analyzed to evaluate the effects of mRNA stability on the target gene expression. Based on the poly A signal of 3' -untranslated region (UTR), nucleotide sequences of putative cytoplasmic polyadenylation elements (CPEs) and downstream elements (DSEs: U-rich, G-rich, GU-rich) were analyzed and used to construct 15 luciferase reporter vectors. Each vector was transfected to HC11 and porcine mammary gland cell (PMGC) and measured for dual luciferase expression levels after 48 hours of incubation. Luciferase expression was significantly higher in construct #6 (with CPE 2, 3 and DSE 1 of exon 9) and #11 (with CPE 2, 3 and DSE 1, 2 and 3 of exon 9) than construct #1 in the PMGC. These results suggest that expression of target genes in PMGC may be effectively expressed by using the construct #6 and #11 on production of transgenic pig.