Efficient and quantitative high-throughput tRNA sequencing

Efficient and quantitative high-throughput transfer RNA sequencing

Despite its biological importance, transfer RNA (tRNA) could not be adequately sequenced by standard methods due to abundant post-transcriptional modifications and stable structure, which interfere with cDNA synthesis. We achieve efficient and quantitative tRNA sequencing using engineered demethylases to remove base methylations and a highly processive thermostable group II intron reverse trans...

Determination of tRNA aminoacylation levels by high-throughput sequencing

Transfer RNA (tRNA) decodes mRNA codons when aminoacylated (charged) with an amino acid at its 3' end. Charged tRNAs turn over rapidly in cells, and variations in charged tRNA fractions are known to be a useful parameter in cellular responses to stress. tRNA charging fractions can be measured for individual tRNA species using acid denaturing gels, or comparatively at the genome level using micr...

High-throughput sequencing technologies.

The human genome sequence has profoundly altered our understanding of biology, human diversity, and disease. The path from the first draft sequence to our nascent era of personal genomes and genomic medicine has been made possible only because of the extraordinary advancements in DNA sequencing technologies over the past 10 years. Here, we discuss commonly used high-throughput sequencing platfo...

SnapShot: High-Throughput Sequencing Applications

High throughput sequencing reveals a plethora of small RNAs including tRNA derived fragments in Haloferax volcanii

To define the complete sRNA population of the halophilic archaeon Haloferax volcanii, we employed high throughput sequencing. cDNAs were generated from RNA ranging in size from 17 to 500 nucleotides isolated from cells grown at three different conditions to exponential and stationary phase, respectively. Altogether, 145 intergenic and 45 antisense sRNAs were identified. Comparison of the expres...