Chlamydomonas (Chlorophyceae) colony PCR.
The ease and effectiveness of colony polymerase chain reaction (PCR) has allowed rapid amplification of DNA fragments and screening of large number of colonies of interest including transformants and mutants with genetic manipulations. Here, we evaluated colony PCR in Chlamydomonas. Individual colonies were treated with 10 mM ethylenediaminetetraacetic acid (EDTA) or Chelex-100 and the resulting clear cell lysate was used for PCR reaction. Either genomic DNA or plasmid DNA incorporated into the genome was equally amplified. We found that the Chelex method is superior to EDTA method in certain cases. This colony PCR technique will bypass the tedious process of isolating genomic DNA for PCR reaction and will make it possible for rapid amplification of genomic DNA fragments as well as rapid large-scale screening of transformants.
Cao
Muqing
M
Department of Biological Sciences and Technology, Tsinghua University, Beijing, 100084, People's Republic of China.
Fu
Yu
Y
Guo
Yan
Y
Pan
Junmin
J
D000818:Animals; D002614:Chelating Agents / Q000737:chemistry; D002696:Chlamydomonas / Q000235:genetics*; D016054:DNA, Protozoan / Q000302:isolation & purification*; D004492:Edetic Acid / Q000737:chemistry; D016133:Polymerase Chain Reaction / Q000379:methods*; D011137:Polystyrenes / Q000737:chemistry; D011145:Polyvinyls / Q000737:chemistry
D002614:Chelating Agents; D016054:DNA, Protozoan; D011137:Polystyrenes; D011145:Polyvinyls; C006960:chelex; D004492:Edetic Acid