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Sample GSM6670721 Query DataSets for GSM6670721
Status Public on Aug 16, 2024
Title opossum, 1M, 1
Sample type SRA
 
Source name testis
Organism Monodelphis domestica
Characteristics tissue: testis
age: 1 month
Growth protocol Animals were grown under standard laboratory conditions and euthanized at the indicated developmental time points.
Extracted molecule total RNA
Extraction protocol Bulk RNA-seq libraries were prepared using KAPA mRNA HyperPrep Kit (Roche # 08098123702) and sequenced on an Illumina NovaSeq
Testes from gray short-tailed opossums at Yale or adult (8-16 week old) male C57BL6/J mice purchased from Jackson Laboratories were harvested and dissociated into single-cell suspension for RNA isolation using collagenase and trypsin. Briefly, testes were removed from the scrotum, decapsulated from the tunica albuginea, and incubated in 0.75mg/mL collagenase in Dulbecco’s Modified Eagle Medium (Gibco 11965-09L; Thermo Fisher Scientific, Inc.) at 37oC. DMEM was added to dilute collagenase and samples were centrifuged for 5 minutes at 400 x g at room temperature. Supernatant was discarded and samples were washed in DMEM. Samples were resuspended in 0.05% trypsin-EDTA (Gibco 25300-054; Thermo Fisher Scientific Inc.) with DNAse (1:10,000; Stem Cell Technologies #07900, Vancouver BC, Canada). The reaction was quenched with 10% Cosmic Calf Serum (Sigma-Aldrich #C8056, Burlington, MA, USA) in DMEM (CCS-DMEM) and centrifuged. Samples were resuspended in CCS-DMEM and filtered through a 100-micron filter. Cell concentration was determined by hemocytometer.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Description polyA RNA-seq
Data processing Assembly: ASM229v1
RNA-seq libraries were aligned using HISAT 2.2.1 (Kim et al., 2019) with Ensembl (Yates et al., 2020) release 104 as a reference and -data output type. Output files were converted from SAM format to sorted BAM format using SAMtools-1.12 (Danecek et al., 2021; Li et al., 2009; Murat et al., 2021). Raw count values and FPKM were obtained using StringTie 2.1.4 (Pertea et al., 2016) with the -e and -G options and Ensembl (Yates et al., 2020) ASM229 release 104 or GRCM39 release 104 reference files for opossum and mouse, respectively.
Supplementary files format and content: scRNA-seq: filtered index (.tsv), feature (.tsv), and matrix (.mtx) files
Supplementary files format and content: bulk RNA-seq: tab-delimited text files with FPKM values by gene
 
Submission date Oct 21, 2022
Last update date Aug 16, 2024
Contact name Bluma J Lesch
Organization name Yale University
Department Genetics
Street address 333 Cedar Street
City New Haven
State/province CT
ZIP/Postal code 06525
Country USA
 
Platform ID GPL30856
Series (1)
GSE216343 Gene regulatory divergence in spermatogenesis between mouse and opossum
Relations
BioSample SAMN31408356
SRA SRX17990194

Supplementary file Size Download File type/resource
GSM6670721_Md_22AB2_e_abundance.tab.gz 834.4 Kb (ftp)(http) TAB
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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