Experiment / E1D6AFD06AAV-MPRA / in vivo MPRA

scAAV miniMPRA of mouse brain enhancer candidates at P7

Parallel functional testing identifies enhancers active in early postnatal mouse brain

A 16-element mouse miniMPRA library containing putative enhancer candidates and negative controls was cloned into the 3′ UTR of an Hsp1a minimal-promoter EGFP reporter. The scAAV9(2YF) library was delivered to P0 mouse prefrontal cortex and assayed in P7 brain using four biological replicate RNA/DNA sequencing measurements.

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Perturbation & assay details

Basal / Untreated

The miniMPRA tested 16 long (approximately 900 bp) mouse noncoding sequences selected as putative enhancers or negative controls. Candidate sequences were cloned into the reporter 3′ UTR, packaged in scAAV9(2YF), injected into P0 prefrontal cortex, and collected at P7; previral Maxiprep and four P7 RNA libraries were counted, normalized to library proportions, and summarized as RNA/DNA activity relative to Maxi_prop.

Processed data

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Column dictionary · 28 definitions
element_id
Unique miniMPRA tested-sequence label from the authors.
negative_control
Package label inferred from an element_id beginning with NEG.
orthologous_star408_amplicon_number
Amplicon number of the corresponding human ortholog in the STAR408 library, from the authors' ID field.
chromosome_mm10
Mouse chromosome for the tested element.
start_mm10
Start coordinate of the tested element on mm10.
end_mm10
End coordinate of the tested element on mm10.
length_bp
Inclusive coordinate span calculated as end_mm10 - start_mm10 + 1.
previral_maxiprep_count
Raw count in the previral Maxiprep library (Maxi).
replicate_1_rna_count
Raw RNA-library count for biological replicate S1.
replicate_2_rna_count
Raw RNA-library count for biological replicate S2.
replicate_3_rna_count
Raw RNA-library count for biological replicate S3.
replicate_4_rna_count
Raw RNA-library count for biological replicate S4.
mean_rna_count
Mean raw RNA count across S1-S4.
sd_rna_count
Standard deviation of raw RNA counts across S1-S4.
previral_maxiprep_proportion
Maxi count normalized to the total previral library count.
replicate_1_rna_proportion
S1 RNA count normalized to that library's total count.
replicate_2_rna_proportion
S2 RNA count normalized to that library's total count.
replicate_3_rna_proportion
S3 RNA count normalized to that library's total count.
replicate_4_rna_proportion
S4 RNA count normalized to that library's total count.
mean_rna_proportion
Mean normalized RNA proportion across S1-S4.
sd_rna_proportion
Standard deviation of normalized RNA proportions across S1-S4.
replicate_1_activity
RNA/DNA ratiometric activity for S1 relative to Maxi_prop.
replicate_2_activity
RNA/DNA ratiometric activity for S2 relative to Maxi_prop.
replicate_3_activity
RNA/DNA ratiometric activity for S3 relative to Maxi_prop.
replicate_4_activity
RNA/DNA ratiometric activity for S4 relative to Maxi_prop.
mean_activity
Mean ratiometric activity across S1-S4.
sd_activity
Standard deviation of ratiometric activity across S1-S4.
qc_pass
Package QC retention flag; true means all five library/RNA count thresholds passed.

Quality control

Supplementary file 1 provides all 16 tested elements. Package QC retained an element when the previral Maxiprep count and each of the four RNA replicate counts were at least 200; all 16/16 passed (minimum Maxiprep count 709 and minimum RNA replicate count 829).

Curation notes

This is the authors' small-scale in vivo proof-of-principle MPRA, distinct from the 408-element human STAR408 screen. Negative-control status is inferred only from labels beginning with NEG; all other source labels are retained without biological reclassification. Coordinates and the orthologous STAR408 number are taken directly from supplementary file 1.

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