run_metadata: 34802

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rowid	run.accession	experiment.accession	sample.accession	study.accession	bioproject	study.title	study.alias	study.type	study.abstract	study.attributes	study.PMIDs	sample.description	sample.title	sample.alias	sample.centername	sample.attributes	GEOsample.title	GEOsample.dataprocessing	GEOsample.source	GEOsample.treatmentprotocol	GEOsample.extractprotocol	GEOsample.growthprotocol	GEOsample.characteristics	GEOsample.accession	experiment.title	experiment.alias	experiment.library_name	experiment.design_description	experiment.library_construction_protocol	experiment.attributes	experiment.library_strategy	experiment.library_source	experiment.library_selection	experiment.library_layout	experiment.platform	experiment.instrument_model	experiment.spot_descriptor	experiment.study_ref	run.title	run.attributes	run.filename	run.semantic_name	run.total_bases	run.total_spots	run.alias	run.read_lengths	run.base_counts	run.r1_length	run.r2_length	run.r3_length	run.r4_length	run.Acount	run.Ccount	run.Gcount	run.Tcount	run.Ncount	run.experiment	run.pool_member	submission.accession	submission.srasource	submission.bioprojectsource	seqdetective.n_mates	seqdetective.mapping_rate.mate1	seqdetective.mapping_rate.mate2	seqdetective.nofeature_rate.mate1	seqdetective.nofeature_rate.mate2	seqdetective.sparsity.mate1	seqdetective.sparsity.mate2	seqdetective.pos_strand_rate.mate1	seqdetective.pos_strand_rate.mate2	seqdetective.readlen.mate1	seqdetective.readlen.mate2	seqdetective.judgement.mate1	seqdetective.judgement.mate2	seqdetective.judgement.reason	platform_family	instrument_generation	read_bias	selection_class	prep_kit	sc_or_bulk	tech_class	technology	tech_variant	submission.bioprojectsource.country	earliest_date	devstage_curation	devstage_curation_coarse	tissue_curation	tissue_curation_coarse
34802	SRR32232919	SRX27572934	SRS23984587	SRP561592	PRJNA1219235	Neurobehavioral phenotypes in the pan neuronal GAL4 zebrafish line Tgelavl3:KalTA4	GSE288775	Transcriptome Analysis	The Gal4/UAS system is used across model organisms to overexpress target genes in precise cell types and relies on generating transgenic Gal4 driver lines. In zebrafish the Tgelavl3:KalTA4 HuC line drives robust expression in neurons. We observed an increased prevalence of swim bladder defects in Tgelavl3:KalTA4 zebrafish larvae compared to wildtype siblings which prompted us to investigate whether transgenic larvae display additional neurobehavioral phenotypes. Tgelavl3:KalTA4 larvae showed alterations in brain activity brain morphology and behavior including increased hindbrain size and reduced activity of the cerebellum. Bulk RNA seq analysis revealed massive dysregulation of the transcriptome and suggested an increased ratio of neuronal progenitor cells compared to differentiated neurons. To understand whether these phenotypes derive from Gal4 toxicity or from positional effects related to transgenesis we used economical low pass whole genome sequencing to map the Tol2 mediated insertion site to chromosome eight. Reduced expression of the neighboring gene gadd45ga a known cell cycle regulator is consistent with increased proliferation and suggests a role for positional effects. Challenges with creating alternative pan neuronal lines include the length of the elavl3 HuC promoter 9 kb and random insertion using traditional transgenesis methods. To facilitate the generation of alternative lines we cloned five neuronal promoters atp6v0cb elavl3 rtn1a sncb and stmn1b ranging from 1.7 kb to 4.3 kb and created KalTA4 lines using Tol2 and the phiC31 integrase based pIGLET system. Our study highlights the importance of using appropriate genetic controls and establishes a roadmap for identifying positional effects in new transgenic lines. Overall design: Tgelavl3:KalTA4 zebrafish were crossed with Tg4xUAS hmgn6 zebrafish to produce wildtype elavl3:KalTA4 4xUAS hmgn6 and elavl3:KalTA4; 4xUAS hmgn6 offspring. At 6 dpf heads were collected for sci RNA seq.				Well P1 09E	GSM8775363		source name:6 dpf head\|tissue:6 dpf head\|well:P1 E09\|rtindex:TCGCTAAGCA\|genotype:GAL4 replicate3\|geo loc name:missing\|collection date:missing	Well P1 09E	Demultiplexing trimming alignment filtering and gene count processing were performed on the UMass Chan High Performance Computing cluster using scripts available from https://github.com/bethmartin/sci RNA seq3 pipeline/tree/master. Reads were aligned to GRCz11 release 104 using the Zebrafish Transcriptome Annotation version 4.3.2 Lawson et al. 2020. Assembly: GRCz11 Supplementary files format and content: RDS containing Seurat object with cell level features counts and barcodes Library strategy: sci RNA seq	6 dpf head		tiny sci sci RNA seq protocol Three heads per biological replicate and three biological replicates per genotype were pooled in 100 µl lysis buffer B in DNA LowBind tubes Eppendorf 002431021. Heads were homogenized with a tissue homogenizer at 1 second intervals for 30 45 seconds until no clumps remained. Following fixation with ice cold methanol and dithiobis succinimidyl propionate DSP Thermo Fisher 22586 nuclei were resuspended in sucrose PBS TritonX MgCl2 SPBSTM and sonicated for 12 seconds on low at 4°C using a Bioruptor® Pico Diagenode. Nuclei were resuspended in SPBSTM with dNTPs and 5 ul of each of the twelve samples was pipetted into each well of one column of a twin.tec LoBind PCR plate Eppendorf 0030129512. Reverse transcription ligation final distribution second strand synthesis protease digestion tagmentation PCR amplification and purification were performed as described except that tagmentation was 10 minutes instead of 5 minutes Martin et al. 2023. Approximately 1000 nuclei were added to each well during the final distribution step. Plate 1 primer sequences were used for reverse transcription ligation and PCR. The library was sequenced on a NextSeq500 by the UAB Heflin Center for Genomic Science Core Laboratories using a 75 cycle kit and 34 cycles for Read1 10 cycles for Index and 48 cycles for Read2.	Sci RNA seq was conducted following the “Tiny Sci” protocol Martin et al. 2023. Briefly Tgelavl3:KalTA4 fish were crossed with Tg4xUAS hmgn6 fish to produce wildtype elavl3:KalTA4 4xUAS hmgn6 and elavl3:KalTA4; 4xUAS hmgn6 offspring. At 6 dpf larvae were anesthetized with MS 222 and heads were removed and frozen on dry ice. The remaining body was saved for genotyping.	tissue:6 dpf head\|well:P1 E09\|rtindex:TCGCTAAGCA\|genotype:GAL4 replicate3	GSM8775363	GSM8775363: Well P1 09E; Danio rerio; OTHER	GSM8775363 r1	GSM8775363	1	tiny sci sci RNA seq protocol Three heads per biological replicate and three biological replicates per genotype were pooled in 100 µl lysis buffer B in DNA LowBind tubes Eppendorf 002431021. Heads were homogenized with a tissue homogenizer at 1 second intervals for 30 45 seconds until no clumps remained. Following fixation with ice cold methanol and dithiobis succinimidyl propionate DSP Thermo Fisher 22586 nuclei were resuspended in sucrose PBS TritonX MgCl2 SPBSTM and sonicated for 12 seconds on low at 4°C using a Bioruptor® Pico Diagenode. Nuclei were resuspended in SPBSTM with dNTPs and 5 ul of each of the twelve samples was pipetted into each well of one column of a twin.tec LoBind PCR plate Eppendorf 0030129512. Reverse transcription ligation final distribution second strand synthesis protease digestion tagmentation PCR amplification and purification were performed as described except that tagmentation was 10 minutes instead of 5 minutes Martin et al. 2023. Approximately 1000 nuclei were added to each well during the final distribution step. Plate 1 primer sequences were used for reverse transcription ligation and PCR. The library was sequenced on a NextSeq500 by the UAB Heflin Center for Genomic Science Core Laboratories using a 75 cycle kit and 34 cycles for Read1 10 cycles for Index and 48 cycles for Read2.		OTHER	TRANSCRIPTOMIC	other	PAIRED	ILLUMINA	NextSeq 500		SRP561592		loader:fastq load.py	P1-09E.R1.fastq.gz P1-09E.R2.fastq.gz P1-09E_S69_I1_001.fastq.gz	fastq fastq fastq	24184408.0	262874.0	GSM8775363 r1	0:34 1:48 2:10	A:3491757;C:1565663;G:4816947;T:2727737;N:15848	34	48	10		3491757	1565663	4816947	2727737	15848	SRX27572934	SRS23984587	SRA2070568	UMass Chan	UMass Chan												B		usable mapping rate	illumina	nextseq	unknown	other	unknown	sc	single_cell_plate	scirnaseq		United States	2025-02-04	Larval	Larval	Head	Nervous System