Random-end N25 5′UTR polysome-profiling MPRA in HEK293T
Optimizing 5’UTRs for mRNA-delivered gene editing using deep learningA HEK293T IVT EGFP reporter library with a 25-nt fully randomized 5′UTR and only the T7-compatible 5′ guanine prefix was profiled in two biological replicates. UTR-level MRL values were combined with replicate-total UMI coverage after polysome fractionation.
Processed tables are specific to each experiment. Column names, units, measurements, and table structure are not standardized across the database. Check this experiment’s column definitions and quality-control notes before comparing or combining data.
Perturbation & assay details
Basal / Untreated
The random-end construct removed the fixed 25-nt prefix and used a 25-nt variable 5′UTR downstream of the T7-derived guanine prefix, followed by EGFP and the reporter 3′ UTR. HEK293T cells were transfected and subjected to cycloheximide-stabilized sucrose-gradient polysome profiling. Reverse transcription/template switching introduced UMIs; fraction-specific libraries were sequenced and MRL was calculated from normalized UMI counts.
Processed data
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Filters apply to this table only. The CSV download contains the complete processed table; filtered rows are available through the API.
Column dictionary · 13 definitions
- element_id
- Stable package identifier assigned in lexicographic UTR-sequence order after QC.
- utr_sequence
- The variable 25-nt random-end 5′UTR sequence represented by the reporter library.
- utr_length
- Length of the packaged variable UTR sequence in nucleotides.
- replicate_count
- Number of biological replicates with a finite MRL value for the sequence.
- total_umi_rep1
- Sum of processed UMI counts across all polysome fractions in random-end N25 replicate 1.
- mrl_rep1
- Author-calculated mean ribosome load for random-end N25 replicate 1.
- total_umi_rep2
- Sum of processed UMI counts across all polysome fractions in random-end N25 replicate 2.
- mrl_rep2
- Author-calculated mean ribosome load for random-end N25 replicate 2.
- mrl_mean
- Arithmetic mean of the two replicate MRL values.
- mrl_sd
- Sample standard deviation of the two replicate MRL values.
- mrl_weighted_mean
- Total-UMI-weighted mean MRL across the two replicates, matching the paper’s replicate-combination calculation.
- total_umi_sum
- Sum of total UMI counts across the two biological replicates; the ≥100-read QC threshold is applied to this field.
- total_umi_min
- Minimum total UMI count among the two replicate observations.
Quality control
Following the authors’ replicate-combination workflow, missing replicate observations were treated as zero coverage, total UMI counts were summed across the two biological replicates, and sequences with summed coverage ≥100 were retained. The package additionally required an exact 25-nt A/C/G/T sequence and finite source MRL values. The GEO files yielded 168,279 retained UTRs; lower-coverage rows were excluded.
Curation notes
The source files are GSE232927_processed_random_end_hek293t_N25_r1.csv.gz and _r2.csv.gz. The publication and supplementary material describe approximately 168k high-coverage sequences; the exact current GEO files produce 168,279 after the documented summed-coverage filter. HEK293T was resolved to Cellosaurus CVCL:0063.