|
| Status |
Public on Jun 15, 2017 |
| Title |
Determination of tRNA aminoacylation levels by high throughput sequencing |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
Here we develop a high throughput sequencing method that enables accurate determination of charged tRNA fractions at single base resolution (Charged tRNA-seq). Our method takes advantage of the recently developed DM-tRNA-seq method, but includes additional chemical steps that specifically remove the 3'A residue in the uncharged tRNA. Charging fraction is obtained by counting the fraction of A-ending reads versus A+C-ending reads for each tRNA species. In HEK293T cells, most cytosolic tRNAs are charged at >90% levels, whereas tRNASer and tRNAThr are generally charged at lower levels. These low charging levels were validated using acid denaturing gels. Our method should be widely applicable for investigations of tRNA charging as a parameter for biological regulation.
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| |
|
| Overall design |
3 biological replicates of tRNA from HEK293T cells are analyzed; each biological replicate is also demethylase treated/untreated
|
| |
|
| Contributor(s) |
Evans M, Clark W, Zheng G, Pan T |
| Citation(s) |
28586482 |
| |
| Submission date |
Mar 31, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Tao Pan |
| E-mail(s) |
taopan@uchicago.edu
|
| Phone |
(773) 702-4179
|
| Organization name |
University of Chicago
|
| Department |
Biochemistry and Molecular Biology
|
| Street address |
929 E. 57th Street
|
| City |
Chicago |
| State/province |
Illinois |
| ZIP/Postal code |
60637 |
| Country |
USA |
| |
|
| Platforms (1) |
| GPL11154 |
Illumina HiSeq 2000 (Homo sapiens) |
|
| Samples (6)
|
| GSM2560256 |
HEK293T AlkB demethylase-untreated one-pot charging; replicate 1 |
| GSM2560257 |
HEK293T AlkB demethylase-untreated one-pot charging; replicate 2 |
| GSM2560258 |
HEK293T AlkB demethylase-untreated one-pot charging; replicate 3 |
| GSM2560259 |
HEK293T AlkB demethylase-treated one-pot charging; replicate 1 |
| GSM2560260 |
HEK293T AlkB demethylase-treated one-pot charging; replicate 2 |
| GSM2560261 |
HEK293T AlkB demethylase-treated one-pot charging; replicate 3 |
|
| Relations |
| BioProject |
PRJNA381134 |
| SRA |
SRP102804 |
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