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@article{720756, author = {Stoneberg Holt, Sierra Dawn and Horová, Lucie and Bureš, Petr}, article_location = {Vienna}, article_number = {2}, keywords = {chemical cleavage of mismatch (CCM); cpDNA; heteroduplex analysis (HA); intraspecific sequence studies; phylogeography; SYBR Green II staining}, language = {eng}, issn = {0040-0262}, journal = {Taxon}, title = {Experimental design in intraspecific organelle DNA sequence studies II: Haplotype detection by Chemical Cleavage of Mismatch (CCM) using SYBR Green II staining}, volume = {56}, year = {2007} }
TY - JOUR ID - 720756 AU - Stoneberg Holt, Sierra Dawn - Horová, Lucie - Bureš, Petr PY - 2007 TI - Experimental design in intraspecific organelle DNA sequence studies II: Haplotype detection by Chemical Cleavage of Mismatch (CCM) using SYBR Green II staining JF - Taxon VL - 56 IS - 2 SP - 509-515 EP - 509-515 PB - International Assoc. for Plant Taxonomy SN - 00400262 KW - chemical cleavage of mismatch (CCM) KW - cpDNA KW - heteroduplex analysis (HA) KW - intraspecific sequence studies KW - phylogeography KW - SYBR Green II staining N2 - Studies of organelle DNA sequence at low taxonomic levels present a researcher with specific experimental design challenges. Such studies require sampling numerous individuals, many of which share identical haplotypes. The process is often limited by the time and money required for DNA sequencing. A number of screening techniques have been devised to combat this limitation, but many have not found wide-spread use in botany. An ideal screening technique would be fast, easy, safe, inexpensive, detect 100% of mutations, be suitable for DNA fragments of about 1 kb, and not require complex equipment. This paper compares the heteroduplex analysis (HA) and chemical cleavage of mismatch (CCM) methods for detecting known point mutations and a deletion in a 1 kb region of non-coding cpDNA from the Poa pratensis agg. A new CCM visualization method, staining with SYBR Green II, was tested. CCM is a powerful tool for discovering different haplotypes in DNA sequence studies at low taxonomic levels. It is quicker and less expensive than sequencing each sample. Compared to HA, CCM is much more sensitive and delivers results in a shorter time, although it is more expensive and considerably more labor intensive. Staining with SYBR Green II allows CCM to be effectively implemented in laboratories with limited access to automated sequencing equipment. ER -
STONEBERG HOLT, Sierra Dawn, Lucie HOROVÁ a Petr BUREŠ. Experimental design in intraspecific organelle DNA sequence studies II: Haplotype detection by Chemical Cleavage of Mismatch (CCM) using SYBR Green II staining. \textit{Taxon}. Vienna: International Assoc. for Plant Taxonomy, 2007, roč.~56, č.~2, s.~509-515. ISSN~0040-0262.
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