Induction of male neogametogenesis in a three-dimensional microenvironment supporting successful fertilization and proper embryo development
Data files
May 13, 2026 version files 1.44 GB
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Control_Sample1.fastq.gz
34.76 MB
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Control_Sample10.fastq.gz
32.18 MB
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Control_Sample2.fastq.gz
34.52 MB
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Control_Sample3.fastq.gz
40.40 MB
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Control_Sample4.fastq.gz
37.85 MB
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Control_Sample5.fastq.gz
36.07 MB
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Control_Sample6.fastq.gz
36.22 MB
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Control_Sample7.fastq.gz
36.40 MB
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Control_Sample8.fastq.gz
35.83 MB
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Control_Sample9.fastq.gz
37.37 MB
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D22_Sample1.fastq.gz
38.27 MB
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D22_Sample10.fastq.gz
40.26 MB
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D22_Sample2.fastq.gz
40.16 MB
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D22_Sample3.fastq.gz
32.40 MB
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D22_Sample4.fastq.gz
39.35 MB
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D22_Sample5.fastq.gz
38.88 MB
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D22_Sample6.fastq.gz
33.87 MB
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D22_Sample7.fastq.gz
33.65 MB
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D22_Sample8.fastq.gz
34.48 MB
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D22_Sample9.fastq.gz
31.18 MB
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D29_Sample1.fastq.gz
38.48 MB
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D29_Sample10.fastq.gz
31.97 MB
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D29_Sample2.fastq.gz
39.99 MB
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D29_Sample3.fastq.gz
32.98 MB
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D29_Sample4.fastq.gz
37.92 MB
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D29_Sample5.fastq.gz
33.47 MB
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D29_Sample6.fastq.gz
40.08 MB
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D29_Sample7.fastq.gz
35.52 MB
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D29_Sample8.fastq.gz
37.93 MB
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D29_Sample9.fastq.gz
32.97 MB
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D36_Sample1.fastq.gz
36.87 MB
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D36_Sample10.fastq.gz
35.60 MB
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D36_Sample2.fastq.gz
31.91 MB
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D36_Sample3.fastq.gz
37.02 MB
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D36_Sample4.fastq.gz
33.14 MB
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D36_Sample5.fastq.gz
31.43 MB
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D36_Sample6.fastq.gz
30.14 MB
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D36_Sample7.fastq.gz
38.77 MB
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D36_Sample8.fastq.gz
44.44 MB
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D36_Sample9.fastq.gz
37.27 MB
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README.md
5.85 KB
Abstract
In regenerative medicine, several attempts have been made to produce functional de novo gametes from mouse embryonic stem cells by utilizing 3D culture systems. We attempted to perform neogametogensis in a novel 3D niche to generate neo-gametes ready to be used for insemination. Mouse ESCs were initially cultured on a gelatin-coated 6-well plate with a monolayer of fibroblasts before being encapsulated in sodium alginate spheres. These spheres were then immersed in specially formulated EpiLC followed by PGCLC medium to promote differentiation into germ-like cells. Over the course of differentiation, immunofluorescence analysis revealed consistent expression patterns of spermatogenic markers. Cells were assessed for DAZL (early germ cell marker) VASA (spermatocyte), BOULE (post-meiotic) and acrosin (spermatid). The differentiated cells were then injected into oocytes and activated by calcium ionophore. Embryo development was monitored via time-lapse microscopy. Spherified neogametes on D22, 29, and 36 achieved fertilization rates of 61.1%, 82.7%, and 80.0%, respectively, and blastulation rates of 20.5%, 36.0%, 26.3%, respectively. Controls exhibited fertilization rates of 89.1% and blastulation rates of 76.3%. When embryo morphokinetics were taken in consideration, D29 embryos reached compaction at 64h and blastulation at 77h, mirroring closely the control’s timing of 63h and 77h, respectively. D22 embryos displayed the most delayed embryo development, of which compaction occurred at 68h and blastulation at 106h. Despite normal fertilization and successful blastulation, the efficiency rate remained below optimal levels. Nevertheless, our system could produce viable offspring, demonstrating that replicating the three-dimensional seminiferous tubule environment is crucial for generating artificial gametes. This eliminates the need for allo-/xeno-genic transplantation in experimental animals.
Dataset DOI: 10.5061/dryad.j0zpc86wh
Description of the data and file structure
Ploidy study on the conceptuses
Files and variables
File: Control_Sample1.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample2.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample3.fastq.gz
Description: Aneuploid embryo generated from injection of natural spermatozoon
File: Control_Sample4.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample5.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample6.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample7.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample8.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample9.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: Control_Sample10.fastq.gz
Description: Euploid embryo generated from injection of natural spermatozoon
File: D22_Sample1.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample2.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample3.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample4.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample5.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample6.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample7.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample8.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample9.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D22_Sample10.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D22 sphere
File: D29_Sample1.fastq.gz
Description: Euploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample2.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample3.fastq.gz
Description: Euploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample4.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample5.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample6.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample7.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample8.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample9.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D29_Sample10.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D29 sphere
File: D36_Sample1.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample2.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample3.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample4.fastq.gz
Description: Euploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample5.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample6.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample7.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample8.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample9.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
File: D36_Sample10.fastq.gz
Description: Aneuploid embryo generated from injection of neogamete extracted from D36 sphere
Code/software
Command Line Tool downloaded and installed through XCode, available from Apple (Mac)
Alternatively, a programming library such as BioPython 1.87 from Python can be used
Access information
Other publicly accessible locations of the data:
- N/A
Data was derived from the following sources:
- N/A
A blastomere biopsy was performed on some embryos at the 8-cell stage for aneuploidy assessment. Whole genome amplification was performed on the samples using the SurePlex DNA Amplification kit (Illumina, San Diego, CA, USA), and next generation sequencing was used to identify chromosomal gain(s) and/or loss(es), as well as to detect mosaicism.