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Image Search Results

Journal: International Journal of Molecular Sciences
Article Title: Human Globozoospermia-Related Gene Spata16 Is Required for Sperm Formation Revealed by CRISPR/Cas9-Mediated Mouse Models
doi: 10.3390/ijms18102208
Figure Lengend Snippet: Targeted point mutation (851G→A, R284Q) of Spata16 by CRISPR/Cas9. ( A ) Targeting scheme of the point mutation (851G→A, R284Q) in the fourth exon of mouse Spata16 . Spata16 consists of eleven exons. The point mutation was introduced into mouse ES cells using the HDR donor plasmid with 0.5 kb homology arms. Blue indicates the sgRNA sequence. Red indicates a G→A mutation at the last nucleotide (851st coding exon) of the fourth exon of Spata16 . Capital and small letters indicate nucleotides of exons and introns, respectively; ( B ) The genotyping of point-mutant mice by Nci I digestion. The Nci I recognition site (5′-CCCGG-3′) was disrupted due to the G→A mutation of the fourth exon. In wild-type mice, two bands (1372 bp and 1569 bp) were detected after Nci I digestion; ( C ) Direct sequencing of PCR products in Spata16 pm/pm mice. The G→A mutation at the last nucleotide (851st coding exon) of the fourth exon is indicated by the orange square; ( D ) RT-PCR analysis of a testis in Spata16 pm/pm mice. The 498-bp band was amplified from wild-type and Spata16 pm/pm mice. A point mutation causing an inappropriate splicing was not found in Spata16 pm/pm mouse testes; ( E ) Representative testicular histology sections stained with hematoxylin and eosin. Spermatogenesis in Spata16 pm/pm mice looked normal compared with that in Spata16 pm/wt mice. Scale bars: 100 μm; ( F ) Cauda epididymal spermatozoa from Spata16 pm/wt and Spata16 pm/pm mice. There were no sperm morphology differences between Spata16 pm/wt and Spata16 pm/pm mice. Spata16 pm/pm males were fertile. Scale bars: 50 μm.
Article Snippet: Spata16 point mutant mice were genotyped by
Techniques: Mutagenesis, CRISPR, Plasmid Preparation, Sequencing, Reverse Transcription Polymerase Chain Reaction, Amplification, Staining