Evaluation of a method for high yield purification of largely intact mitochondrial DNA from human placentae
Genet. mol. res. (Online)
;
2(2): 178-184, Jun. 2003.
Artigo
em Inglês
| LILACS
| ID: lil-417612
RESUMO
We developed, and quantitatively and qualitatively evaluated an easily reproducible method for high yield purification of mitochondrial DNA (mtDNA) from human placentae by mechanical tissue disruption, differential centrifugation of mitochondria, enzymatic digestion, phenol extraction and ethanol precipitation. Average mtDNA yields were 2.5 microg/g tissue (without an RNAse treatment step) and 1.5 microg/g tissue (with an RNAse treatment step). This mtDNA migrated as a 16.5-kb isolated band in agarose gels; it yielded fragments of expected sizes after digestion with restriction enzymes; it successfully served as a template in long PCR for amplification of mtDNA sequences, and hybridized to an mtDNA probe in a predictable fashion. MtDNA yields of this method were 10-fold higher than those of previously reported ones for mtDNA purification from freshly obtained human cells and tissues, with the advantage that more placental tissue can be obtained for mtDNA purification than other types of tissue, at lower cost, and with minimal or no ethical issues
Texto completo:
DisponíveL
Índice:
LILACS (Américas)
Assunto principal:
Placenta
/
DNA Mitocondrial
Limite:
Humanos
Idioma:
Inglês
Revista:
Genet. mol. res. (Online)
Assunto da revista:
Biologia Molecular
/
Genética
Ano de publicação:
2003
Tipo de documento:
Artigo
País de afiliação:
Brasil
Instituição/País de afiliação:
AADM Gene Therapy Center and Genon Ltd/BR
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