Analysis of cell cycle progression and genomic integrity in early lethal knockouts.
Owing to their importance in normal cell division, DNA damage checkpoint and repair genes are often required for the earliest stages of embryzonic development. For example, conventional deletion of ATR, Chk1, Mad2, NBS, Rad50, BRCA1, BRCA2, or Rad51 leads to developmental arrest prior to gastrulation. While prior to arrest the number of cells extant in these embryos is low, procedures allowing rudimentary analysis of cell cycle checkpoints and genome integrity have been developed through culturing blastocysts in vitro. These procedures provide a small number of proliferating cells that can be analyzed for cell cycle progression, G2/M phase checkpoint responses, and gross chromosome abnormalities by mitotic spread preparation. Experiments such as these may help determine the essential functions of these genes in cell proliferation and early embryonic development. It is interesting to note that recently developed methods to introduce single-copy transgenes into one-cell zygotes via lentiviruses may provide a means to generate Cre/lox-conditional cell lines from these conventional knockouts.
Brown
Eric J
EJ
Department of Cancer Biology, Abramson Family Cancer Research Institute, University of Pennsylvania School of Medicine, Philadelphia, USA.
D000818:Animals; D017209:Apoptosis; D001755:Blastocyst / Q000378:metabolism; D015139:Blotting, Southern; D002453:Cell Cycle; D018797:Cell Cycle Proteins; D002455:Cell Division; D002869:Chromosome Aberrations; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D016195:G2 Phase; D018816:Genes, cdc; D005816:Genetic Complementation Test; D005838:Genotype; D006579:Heterozygote; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D008938:Mitosis; D015739:Nocodazole / Q000494:pharmacology; D009841:Oligonucleotides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011494:Protein Kinases
D018797:Cell Cycle Proteins; D009841:Oligonucleotides; D004247:DNA; D011494:Protein Kinases; D015739:Nocodazole