65 research outputs found
A High-Speed Congenic Strategy Using First-Wave Male Germ Cells
BACKGROUND: In laboratory mice and rats, congenic breeding is essential for analyzing the genes of interest on specific genetic backgrounds and for analyzing quantitative trait loci. However, in theory it takes about 3-4 years to achieve a strain carrying about 99% of the recipient genome at the tenth backcrossing (N10). Even with marker-assisted selection, the so-called 'speed congenic strategy', it takes more than a year at N4 or N5. METHODOLOGY/PRINCIPAL FINDINGS: Here we describe a new high-speed congenic system using round spermatids retrieved from immature males (22-25 days of age). We applied the technique to three genetically modified strains of mice: transgenic (TG), knockin (KI) and N-ethyl-N-nitrosourea (ENU)-induced mutants. The donor mice had mixed genetic backgrounds of C57BL/6 (B6):DBA/2 or B6:129 strains. At each generation, males used for backcrossing were selected based on polymorphic marker analysis and their round spermatids were injected into B6 strain oocytes. Backcrossing was repeated until N4 or N5. For the TG and ENU-mutant strains, the N5 generation was achieved on days 188 and 190 and the proportion of B6-homozygous loci was 100% (74 markers) and 97.7% (172/176 markers), respectively. For the KI strain, N4 was achieved on day 151, all the 86 markers being B6-homozygous as early as on day 106 at N3. The carrier males at the final generation were all fertile and propagated the modified genes. Thus, three congenic strains were established through rapid generation turnover between 41 and 44 days. CONCLUSIONS/SIGNIFICANCE: This new high-speed breeding strategy enables us to produce congenic strains within about half a year. It should provide the fastest protocol for precise definition of the phenotypic effects of genes of interest on desired genetic backgrounds
Clinical significance of heparin-binding epidermal growth factor-like growth factor in peritoneal fluid of ovarian cancer
Epidermal growth factor receptor (EGFR) has been implicated in tumour growth and extension of ovarian cancer. Peritoneal fluid in ovarian cancer patients contains various growth factors that can promote tumour growth and extension. In order to investigate the clinical significance of EGFR ligands as activating factors of ovarian cancer, we examined the cell proliferation-promoting activity and the level of EGFR ligands in peritoneal fluid obtained from 99 patients. Proliferation-promoting activity in peritoneal fluid from 63 ovarian cancer patients (OVCA) was much higher than peritoneal fluid from 18 ovarian cyst patients (OVC) and 18 normal ovary patients (NO), and the activity was suppressed only by antibodies against EGFR or heparin-binding epidermal growth factor (HB-EGF). A large difference was observed in the level of EGFR ligands between HB-EGF and TGF-Ξ± or amphiregulin. The concentration of HB-EGF in OVCA significantly increased compared to that in OVC or NO (P<0.01). No significant difference in the concentration of TGF-Ξ± and amphiregulin was found between the OVCA and NO or OVC groups. In peritoneal fluid, HB-EGF is sufficiently elevated to activate cancer cells even at an early stage of OVCA. These results suggested that HB-EGF in peritoneal fluid might play a key role in cell survival and in the proliferation of OVCA
Analysis of Lewis fucosyltransferase genes from the human gastric mucosa of Lewis-positive and -negative individuals
Analysis of Lewis fucosyltransferase genes from the human gastric mucosa of Lewis-positive and -negative individuals
Modification of cell membranes with viral envelopes during fusion of cells with hvj (sendai virus). Iii. Effects of mono- and di- -saccharides on cell fusion and membrane movement of fused cells.
In vitro studies on odontogenic tumors
Ameloblastomas are uncommon benign neoplasms of the jaws. They originate from dental epithelial cells, but they are not capable of mineralizing or forming enamel. The study of these tumors is limited to live tissue collected from patients during scheduled surgery. Ameloblastomas grow slowly in vivo and this property is translated to their behavior in vitro. Here, we describe the methods to culture ameloblastomas in organotypic cultures, as well as to isolate stem/progenitor cells from these tumors
Phorbol ester induces the rapid processing of cell surface heparin-binding EGF-like growth factor: conversion from juxtacrine to paracrine growth factor activity.
Membrane translocation and channel-forming activities of diphtheria toxin are blocked by replacing isoleucine 364 with lysine
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