DOI: 10.5061/dryad.zkh1893kb
title: Single-cell RNA sequencing of the testis of drive and standard Teleopsis dalmanni males

description: Sex-linked meiotic drivers limit the inheritance of the alternate sex
 chromosome in the heterogametic sex, subsequently skewing the ratio of
 males to females in the offspring. They consequently have large impacts on
 genome evolution, adaptation, and the emergence and maintenance of
 sexually selected traits. Detecting the molecular basis of meiotic drive
 is complex. It is affected by both the common occurrence of inversions
 housing a driver as well as the transcriptomic and cell type heterogeneity
 of the gonads. Herein, we utilise DNA-Seq to investigate the genetic basis
 of a sex-linked meiotic driver in the Malaysian stalk-eyed fly, Teleopsis
 dalmanni. These data are generated from wild adult flies from the Gombak
 District of Peninsular Malaysia. 
description: # Single-cell RNA sequencing of the testis of the drive and standard
 stalk-eyed fly.
 [https://doi.org/10.5061/dryad.zkh1893kb](https://doi.org/10.5061/dryad.zkh1893kb) ## Description of the data and file structure **Samples** Flies were reared at University College London from a wild-caught population originating from the Gombak Valley, Malaysia. All flies and larvae were incubated and reared at 25°C and fed on a diet of sweetcorn. To ensure known genotypes of samples, a homozygous drive (SR) population is maintained through a series of crosses as described in [(Presgraves et al., 1997)](https://paperpile.com/c/UqA1L6/3Hym). Eight adult males, four standard (ST) and four with drive (SR), were sacrificed before the dissection of both testes in iced phosphate-buffered saline (PBS). These adults were all virgins, reproductively mature, and reared from egg lays collected on the same day.  ***Tissue collection, dissociation, and single-cell sequencing*** Testis pairs were individually dissociated by incubation in a collagenase-TrypLE lysis solution (10mg/ml collagenase in 10X TrypLE) at 37.5°C for one hour with three sets of mechanical dissociation by triturations of wide then narrow bore Pasteur pipettes. Digestion was inhibited by the addition of iced Schneider’s Serum. The solution was then gently pipetted up and down with a narrow-bore Pasteur pipette before filtering through a 35μm filter pre-rinsed with Schneider’s Serum. The sample was then spun in a swing bucket centrifuge for 5 minutes at 1000xg and 4°C. The supernatant was removed and the pellet resuspended in 50μl of iced PBS with gentle pipetting of a wide-bore pipette. To count cells, 10μl of the suspension was combined with 10μl of trypan blue and placed onto a humidified haemocytometer plate before counting in triplicate.  10X Genomics Chromium transcriptome libraries were generated at the NERC Environmental Omics Facility (NEOF), Liverpo, before sequencing with Illumina NovaSeq using S2 chemistry, aiming for recovery of ~10,000 cells per sample and ~20,000 reads per cell.  ### Files and variables **3-sr1_TAACGCGT_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr1, Lane 002, containing the sample index barcode. **3-sr1_TAACGCGT_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr1, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **3-sr1_TAACGCGT_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr1, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **3-sr1_TAACGCGT_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr1, Lane 001, containing the sample index barcode. **3-sr1_TAACGCGT_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr1, Lane 001, containing the actual cDNA sequence. **3-sr1_TAACGCGT_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr1, Lane 002, containing the actual cDNA sequence. **4-sr2_AATGCCAT_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr2, Lane 001, containing the actual cDNA sequence. **4-sr2_AATGCCAT_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr2, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **4-sr2_AATGCCAT_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr2, Lane 002, containing the sample index barcode. **4-sr2_AATGCCAT_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr2, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **4-sr2_AATGCCAT_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr2, Lane 001, containing the sample index barcode. **4-sr2_AATGCCAT_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr2, Lane 002, containing the actual cDNA sequence. **5-sr3_ACGACTAC_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr3, Lane 001, containing the sample index barcode. **5-sr3_ACGACTAC_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr3, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **5-sr3_ACGACTAC_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr3, Lane 002, containing the sample index barcode. **5-sr3_ACGACTAC_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr3, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **5-sr3_ACGACTAC_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr3, Lane 002, containing the actual cDNA sequence. **5-sr3_ACGACTAC_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr3, Lane 001, containing the actual cDNA sequence. **7-st1_TTGAGAGT_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st1, Lane 001, containing the sample index barcode. **7-st1_TTGAGAGT_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st1, Lane 002, containing the sample index barcode. **7-st1_TTGAGAGT_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st1, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **7-st1_TTGAGAGT_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st1, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **7-st1_TTGAGAGT_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st1, Lane 002, containing the actual cDNA sequence. **7-st1_TTGAGAGT_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st1, Lane 001, containing the actual cDNA sequence. **8-st2_TTGCCCGT_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st2, Lane 001, containing the sample index barcode. **8-st2_TTGCCCGT_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st2, Lane 002, containing the sample index barcode. **8-st2_TTGCCCGT_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st2, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **8-st2_TTGCCCGT_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st2, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **8-st2_TTGCCCGT_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st2, Lane 001, containing the actual cDNA sequence. **8-st2_TTGCCCGT_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st2, Lane 002, containing the actual cDNA sequence. **9-st3_GCGGGTAA_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st3, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **9-st3_GCGGGTAA_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st3, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **9-st3_GCGGGTAA_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st3, Lane 001, containing the actual cDNA sequence. **9-st3_GCGGGTAA_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st3, Lane 001, containing the sample index barcode. **9-st3_GCGGGTAA_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st3, Lane 002, containing the sample index barcode. **9-st3_GCGGGTAA_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st3, Lane 002, containing the actual cDNA sequence. **10-st5_CCTATCCT_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st5, Lane 001, containing the actual cDNA sequence. **10-st5_CCTATCCT_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st5, Lane 002, containing the sample index barcode. **10-st5_CCTATCCT_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st5, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **10-st5_CCTATCCT_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for st5, Lane 002, containing the actual cDNA sequence. **10-st5_CCTATCCT_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for st5, Lane 001, containing the sample index barcode. **10-st5_CCTATCCT_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for st5, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **11-sr5_AGCAAGAA_L001_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr5, Lane 001, containing the sample index barcode. **11-sr5_AGCAAGAA_L002_I1_001.fastq.gz**\ **Description**: Sample index (I1) file for sr5, Lane 002, containing the sample index barcode. **11-sr5_AGCAAGAA_L002_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr5, Lane 002, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **11-sr5_AGCAAGAA_L001_R1_001.fastq.gz**\ **Description**: Cell barcode + UMI (R1) file for sr5, Lane 001, containing the sequence for cell barcode and Unique Molecular Identifier (UMI). **11-sr5_AGCAAGAA_L002_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr5, Lane 002, containing the actual cDNA sequence. **11-sr5_AGCAAGAA_L001_R2_001.fastq.gz**\ **Description**: cDNA sequence (R2) file for sr5, Lane 001, containing the actual cDNA sequence. **DATA DESCRIPTION** **T_dalmanni_sample_data_information.xlsx** **Description**: Spreadsheet containing data and information on gonad samples from the Stalkies project. * Sheet 1, information includes:  * Sample: sample name given to each sample during dissections and used in downstream analyses * Sequencing sample number: sample number given to each sample by the Centre for Genomic Research at Liverpool during sequencing  * Genotype: indicates whether the individual carries a driving X chromosome (Drive) or does not (Standard) * Date sexed: Indicates the date that sex was acknowledged for the individual * Dissociation date: Date that the testis from each sample was dissociated into single cells and processed for sequencing * Original Solution Vol (ul): Volume of cell suspension * Cells/ul: Concentration of cells within the cell suspension * Sheet 2, information includes  * Sample name * Sequencing sample number * Reads: the sequencing reads belonging to the relevant sample, see above in the README for full information per sample * md5sum: md5 code generated to ensure the integrity of the file upon downloading. **QUALITY CONTROL REPORTS** **sr1-5_st1-3_st5_cdna2100_expert_High_Sensitivity_DNA_Assay_DE72905138_2022-07-07_09-43-57.pdf**\ **Description**: High Sensitivity DNA Assay quality control report for cDNA for combined samples dated July 7, 2022. **sr1-5_st1-5_final_lib2100_expert_High_Sensitivity_DNA_Assay_DE72905138_2022-07-07_13-45-15.pdf**\ **Description**: High Sensitivity DNA Assay quality control report for final libraries for combined samples, dated July 7, 2022.