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Titolo:
Direct injection of foreign DNA into mouse testis as a possible in vivo gene transfer system via epididymal spermatozoa
Autore:
Sato, M; Ishikawa, A; Kimura, M;
Indirizzi:
Tokai Univ, Inst Med Sci, Kanagawa 2591193, Japan Tokai Univ Kanagawa Japan 2591193 Inst Med Sci, Kanagawa 2591193, Japan Tokai Univ, Dept Mol Life Sci, Kanagawa 2591100, Japan Tokai Univ Kanagawa Japan 2591100 Mol Life Sci, Kanagawa 2591100, Japan
Titolo Testata:
MOLECULAR REPRODUCTION AND DEVELOPMENT
fascicolo: 1, volume: 61, anno: 2002,
pagine: 49 - 56
SICI:
1040-452X(200201)61:1<49:DIOFDI>2.0.ZU;2-G
Fonte:
ISI
Lingua:
ENG
Soggetto:
TRANSGENIC MICE; CELLS; EXPRESSION; SEQUENCE; EGGS;
Keywords:
electroporation; epididymis; hoechst 33342; in vivo gene transfer; sperm vector; testis; transfection; trypan blue;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
20
Recensione:
Indirizzi per estratti:
Indirizzo: Sato, M Tokai Univ, Inst Med Sci, Kanagawa 2591193, Japan Tokai Univ Kanagawa Japan 2591193 d Sci, Kanagawa 2591193, Japan
Citazione:
M. Sato et al., "Direct injection of foreign DNA into mouse testis as a possible in vivo gene transfer system via epididymal spermatozoa", MOL REPROD, 61(1), 2002, pp. 49-56

Abstract

We have attempted to transfect testicular spermatozoa with plasmid DNA by direct injection into testes to obtain transgenic animals [this technique was thus termed "testis-mediated gene transfer (TMGT)"]. When injected maleswere mated with superovulated females 2 and 3 days after injection, (i) high efficiencies (more than 50%) of gene transmission were achieved in the midgestational F0 fetuses, (ii) the copy number of plasmid DNA in the fetuses was estimated to be less than I copy per diploid cell, and (iii) overt gene expression was not found in these fetuses. These findings suggest the possibility that plasmid DNA introduced into a testis is rapidly transported to the epididymis and then incorporated by epididymal spermatozoa. The purpose of this study was to elucidate the mechanism of TMGT by introducing trypan blue (TB) or Hoechst 33342 directly into testis. We found that TB is transported to the ducts of the caput epididymis via rete testis within I minafter testis injection, and TB reached the corpus and cauda epididymis within 2-4 days after injection. Staining of spermatozoa isolated from any portion of epididymis was observed 4 days after injection of a solution containing Hoechst 33342. Injection of enhanced green fluorescent protein (EGFP) expression vector/liposome complex into testis resulted in transfection of epithelial cells of epididymal ducts facing the lumen, although the transfection efficiency appeared to be low. In vivo electroporation toward the caput epididymis immediately after injection of EGFP expression vector into a testis greatly improved the uptake of foreign DNA by the epididymal epithelial cells. PCR analysis using spermatozoa isolated from corpus and cauda epididymis 4 days after injection of a DNA/liposome complex into testis revealed exogenous DNA in these spermatozoa even after treatment with DNase I. These findings indicate that exogenous DNA introduced into tesits is rapidlytransported to epididymal ducts via the rete testis and efferent ducts, and then incorporated by epithelial cells of epididymis and epididymal spermatozoa. Mol. Reprod. Dev. 61: 49-56, 2002. (C) 2002 Wiley-Liss, Inc.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 19/02/20 alle ore 02:28:32