Single cells

Credit: © Pascal Goetgheluck / Photo Researchers, Inc." /> Credit: © Pascal Goetgheluck / Photo Researchers, Inc. User: Norma Nowak, Roswell Park Cancer Institute, Buffalo, NY Project: Conducting pre-implantation genetic diagnoses (PGD) on blastomere-derived cells. Problem: Cells harvested for PGD are traditionally put into a generic buffer and frozen. Th

Josh P. Roberts
Apr 1, 2008
<figcaption> Credit: © Pascal Goetgheluck / Photo Researchers, Inc.</figcaption>
Credit: © Pascal Goetgheluck / Photo Researchers, Inc.

User:
Norma Nowak, Roswell Park Cancer Institute, Buffalo, NY

Project:
Conducting pre-implantation genetic diagnoses (PGD) on blastomere-derived cells.

Problem:
Cells harvested for PGD are traditionally put into a generic buffer and frozen. That treatment often results in "allele dropout," in which many chromosomal regions are not amenable to amplification.

Solution:
"You've got only one shot if you do PCR directly from single cells," says Nowak. So she puts cells right into a lysis buffer containing proteinase K immediately after isolating them. The buffer immediately begins to digest nucleases and other proteins, protecting and allowing easier access to the DNA.

Nowak uses the buffer that comes with Sigma's GenomePlex® Whole Genome Amplification Kit. She uses the kit to amplify the DNA, prior to performing PCR. This kills two birds...