Medicine and Health Sciences Commons^™

Gonad: Genome-Editing Via Oviductal Nucleic Acids Delivery System: A Novel Microinjection Independent Genome Engineering Method In Mice., Gou Takahashi, Channabasavaiah B. Gurumurthy, Kenta Wada, Hiromi Miura, Masahiro Sato, Masato Ohtsuka

Journal Articles: Genetics, Cell Biology & Anatomy

Microinjection is considered the gold standard technique for delivery of nucleic acids (NAs; transgenes or genome editing tools such as CRISPR/Cas9 systems) into embryos, for creating genetically modified organisms. It requires sophisticated equipment as well as well-trained and highly skilled personnel to perform the micro-injection technique. Here, we describe a novel and simple microinjection-independent technique, called Genome-editing via Oviductal Nucleic Acids Delivery (GONAD). Using GONAD, we show that NAs (e.g., eGFP mRNA or Cas9 mRNA/sgRNAs) can be effectively delivered to pre-implantation embryos within the intact mouse oviduct by a simple electroporation method, and result in the desired genetic modification in …

A Patient With Polymerase E1 Deficiency (Pole1): Clinical Features And Overlap With Dna Breakage/Instability Syndromes., Isabelle Thiffault, Carol Saunders, Janda Jenkins, Nikita Raje, Kristi Canty, Mukta Sharma, Lauren Grote, Holly I. Welsh, Emily Farrow, Greyson Twist, Neil Miller, David Zwick, Lee Zellmer, Stephen F. Kingsmore, Nicole P. Safina

Manuscripts, Articles, Book Chapters and Other Papers

BACKGROUND: Chromosome instability syndromes are a group of inherited conditions associated with chromosomal instability and breakage, often leading to immunodeficiency, growth retardation and increased risk of malignancy.

CASE PRESENTATION: We performed exome sequencing on a girl with a suspected chromosome instability syndrome that manifested as growth retardation, microcephaly, developmental delay, dysmorphic features, poikiloderma, immune deficiency with pancytopenia, and myelodysplasia. She was homozygous for a previously reported splice variant, c.4444 + 3A > G in the POLE1 gene, which encodes the catalytic subunit of DNA polymerase E.

CONCLUSION: This is the second family with POLE1-deficency, with the affected individual demonstrating a more …

One-Step Generation Of Multiple Transgenic Mouse Lines Using An Improved Pronuclear Injection-Based Targeted Transgenesis (I-Pitt)., Masato Ohtsuka, Hiromi Miura, Keiji Mochida, Michiko Hirose, Ayumi Hasegawa, Atsuo Ogura, Ryuta Mizutani, Minoru Kimura, Ayako Isotani, Masahito Ikawa, Masahiro Sato, Channabasavaiah B. Gurumurthy

Journal Articles: Genetics, Cell Biology & Anatomy

BACKGROUND: The pronuclear injection (PI) is the simplest and widely used method to generate transgenic (Tg) mice. Unfortunately, PI-based Tg mice show uncertain transgene expression due to random transgene insertion in the genome, usually with multiple copies. Thus, typically at least three or more Tg lines are produced by injecting over 200 zygotes and the best line/s among them are selected through laborious screening steps. Recently, we developed technologies using Cre-loxP system that allow targeted insertion of single-copy transgene into a predetermined locus through PI. We termed the method as PI-based Targeted Transgenesis (PITT). A similar method using PhiC31-attP/B system …

Systems Level Analysis Of Systemic Sclerosis Shows A Network Of Immune And Profibrotic Pathways Connected With Genetic Polymorphisms, J. Matthew Mahoney, Jaclyn Taroni, Viktor Martyanov, Tammara A. A. Wood, Casey S. Greene, Patricia A. Pioli, Monique E. Hinchcliff, Michael L. Whitfield

Dartmouth Scholarship

Systemic sclerosis (SSc) is a rare systemic autoimmune disease characterized by skin and organ fibrosis. The pathogenesis of SSc and its progression are poorly understood. The SSc intrinsic gene expression subsets (inflammatory, fibroproliferative, normal-like, and limited) are observed in multiple clinical cohorts of patients with SSc. Analysis of longitudinal skin biopsies suggests that a patient's subset assignment is stable over 6-12 months. Genetically, SSc is multi-factorial with many genetic risk loci for SSc generally and for specific clinical manifestations. Here we identify the genes consistently associated with the intrinsic subsets across three independent cohorts, show the relationship between these genes …