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nrStar™ Human tRNA Repertoire PCR Array
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Figure 1. The effects of tRNA repertoire during cell fate determination
Transfer RNAs (tRNAs) are ubiquitous and the most abundant of all small non-coding RNA molecules. As a fundamental component in translation, tRNAs serve as the physical link between the mRNA coding and protein sequences. A wide variety of biological processes, such as cell proliferation, differentiation[1, 2] and apoptosis, are always accompanied with variation of tRNAs levels. Alterations of tRNA repertoire affect cell-fate choices during cell development (Fig. 1). Dysregulated tRNA repertoire can promote tumorigenesis and cancer progression [2, 4-11]. Additionally, various diseases show disruptions to the levels and distributions of tRNAs, such as type 2 diabetes mellitus , Huntington disease , and HIV infection . Studying tRNA repertoire has become an important part of research of biological processes and human diseases.
PCR Arrays are the most reliable tools for analyzing the expression of a focused panel of genes. nrStar™ Human tRNA Repertoire PCR Array profiles 66 PCR-distinguishable nuclear tRNA isodecoders and all mitochondrial tRNA species. The panel covers all anti-codons provided by GtRNAdb and Transfer RNA database.
tRNAs undergo by far the greatest number of and the most chemically diverse post-transcriptional modifications, which badly affect tRNA cDNA synthesis efficiency. Scientists at Arraystar have developed rtStar™ tRNA-optimized First-Strand cDNA Synthesis Kit(AS-FS-004) that efficiently removes the tRNA modifications and greatly improves the cDNA synthesis quality. With the powerful combination of this kit and the PCR array, researchers can obtain a new level of accuracy on the tRNA pool alterations and gain further insight to interpret the proteome and tRNA-derived fragments.
To ensure the data accuracy, the panel includes 3 reference sets for small non-coding RNAs to better quantify and normalize the qPCR data. cDNA synthesis and PCR efficiency are evaluated by using the RNA Spike-in control. Potential genomic DNA contamination is monitored by using the genomic DNA control (GDC). Positive PCR control (PPC) is designed for monitoring PCR efficiency and inter-plate calibration.
Armed with the tRNA repertoire data, gain-of-function [2, 15, 16] and lose-of-function  approaches are useful for follow up studies. Commonly used methods in non-coding RNA studies are readily applicable to tRNA research in-depth.
 Gingold H, Tehler D, Christoffersen NR, Nielsen MM, Asmar F, Kooistra SM, et al. A dual program for translation regulation in cellular proliferation and differentiation. Cell 2014;158:1281-92.
 Pavon-Eternod M, Gomes S, Rosner MR, Pan T. Overexpression of initiator methionine tRNA leads to global reprogramming of tRNA expression and increased proliferation in human epithelial cells. Rna 2013;19:461-6.
 Mei Y, Stonestrom A, Hou YM, Yang X. Apoptotic regulation and tRNA. Protein & cell 2010;1:795-801.
 Berns A. A tRNA with oncogenic capacity. Cell 2008;133:29-30.
 Waldman YY, Tuller T, Sharan R, Ruppin E. TP53 cancerous mutations exhibit selection for translation efficiency. Cancer research 2009;69:8807-13.
 Kushner JP, Boll D, Quagliana J, Dickman S. Elevated methionine-tRNA synthetase activity in human colon cancer. Proceedings of the Society for Experimental Biology and Medicine Society for Experimental Biology and Medicine 1976;153:273-6.
 Marshall L, Kenneth NS, White RJ. Elevated tRNA(iMet) synthesis can drive cell proliferation and oncogenic transformation. Cell 2008;133:78-89.
 Pavon-Eternod M, Gomes S, Geslain R, Dai Q, Rosner MR, Pan T. tRNA over-expression in breast cancer and functional consequences. Nucleic acids research 2009;37:7268-80.
 Zhou Y, Goodenbour JM, Godley LA, Wickrema A, Pan T. High levels of tRNA abundance and alteration of tRNA charging by bortezomib in multiple myeloma. Biochemical and biophysical research communications 2009;385:160-4.
 Begley U, Sosa MS, Avivar-Valderas A, Patil A, Endres L, Estrada Y, et al. A human tRNA methyltransferase 9-like protein prevents tumour growth by regulating LIN9 and HIF1-alpha. EMBO molecular medicine 2013;5:366-83.
 Goodarzi H, Nguyen HC, Zhang S, Dill BD, Molina H, Tavazoie SF. Modulated Expression of Specific tRNAs Drives Gene Expression and Cancer Progression. Cell 2016;165:1416-27.
 Krokowski D, Han J, Saikia M, Majumder M, Yuan CL, Guan BJ, et al. A self-defeating anabolic program leads to beta-cell apoptosis in endoplasmic reticulum stress-induced diabetes via regulation of amino acid flux. The Journal of biological chemistry 2013;288:17202-13.
 Girstmair H, Saffert P, Rode S, Czech A, Holland G, Bannert N, et al. Depletion of cognate charged transfer RNA causes translational frameshifting within the expanded CAG stretch in huntingtin. Cell reports 2013;3:148-59.
 van Weringh A, Ragonnet-Cronin M, Pranckeviciene E, Pavon-Eternod M, Kleiman L, Xia X. HIV-1 modulates the tRNA pool to improve translation efficiency. Molecular biology and evolution 2011;28:1827-34.
 Gong M, Gong F, Yanofsky C. Overexpression of tnaC of Escherichia coli inhibits growth by depleting tRNA2Pro availability. Journal of bacteriology 2006;188:1892-8.
 Yona AH, Bloom-Ackermann Z, Frumkin I, Hanson-Smith V, Charpak-Amikam Y, Feng Q, et al. tRNA genes rapidly change in evolution to meet novel translational demands. eLife 2013;2:e01339.
 Fu G, Xu T, Shi Y, Wei N, Yang XL. tRNA-controlled nuclear import of a human tRNA synthetase. The Journal of biological chemistry 2012;287:9330-4.
• Profile nuclear and mitochondrial tRNAs of all anti-codons in the GtRNAdb and tRNAdb databases
• Profile all tRNA isodecoders distinguishable by real-time PCR
• Fully experimentally validated primers across numerous tissues and cell lines
• Easy-to-use convenient panel to obtain results in hours
rtStar™ tRNA-optimized First-Strand cDNA Synthesis Kit
The highly efficient system for cDNA synthesis from tRNA
tRNAs undergo by far the greatest number of and the most chemically diverse post-transcriptional modifications, which badly affect tRNA cDNA synthesis efficiency. Methylation, one of the most prevalent modifications, is the main obstacle that hinder transcriptional elongation and thus leads to noneffective cDNA conversion and qPCR. By using an excellent RNA demethylase, Arraystar developed rtStar™ tRNA-optimized First-Strand cDNA Synthesis Kit (Cat# AS-FS-004), a highly efficient system that removes methylations on the RNA to obtain a new level of accuracy on the tRNA detection.
Figure 1. tRNA demethylation treatment dramatically increased PCR assay performance. A. tRNA levels in 3 pancreatic carcinomas and 3 matched para-carcinoma RNA samples with or without the RNA demethylation treatment. Error bars indicate SEM (n=3). De: demethylation, Un: untreated with demethyation. B. Principle component analysis (PCA) analysis of tRNA profiles for the pancreatic carcinoma and para-carcinoma tissues. The Cancer (Demethylation) and Para-carcinoma (Demethylation) samples are well separated, whereas the Cancer and Para-carcinoma samples without the demethylation treatment are indistinguishable.