Genetic fingerprinting of grape plant (Vitis vinifera) using random amplified polymorphic DNA (RAPD) analysis and dynamic size-sieving capillary electrophoresis.
Dynamic size-sieving capillary electrophoresis with laser-induced fluorescence detection (DSCE-LIF) was combined with random amplified polymorphic DNA (RAPD) analysis to demonstrate the feasibility of the genetic analysis of grape plant varieties and clones within a variety. Parameters of the genomic DNA extraction process, as well as those of the RAPD analysis, were optimized specifically for this application. Polymorphic DNA fragments were generated for four different grape plant varieties including Cabernet Franc, Cabernet Sauvignon, Merlot, and Chardonnay. Relative to slab gel electrophoresis (SGE) with ethidium bromide staining, DSCE-LIF provided superior separation efficiency and detection limits in the analysis of DNA polymorphic bands. Optimal DSCE-LIF analyses were achieved using a 10-fold RAPD sample dilution, hydrodynamic sample injection, and 100 ng/mL of YO-PRO-1 DNA intercalator in the dynamic size-sieving buffer solution. In addition, the reproducibility of both the DSCE-LIF and RAPD analyses were demonstrated.
Siles
B A
BA
The College of William and Mary, Department of Chemistry, P.O. Box 8795, Williamsburg, Virginia 23187-8795, USA. bsiles@du.edu
O'Neil
K A
KA
Fox
M A
MA
Anderson
D E
DE
Kuntz
A F
AF
Ranganath
S C
SC
Morris
A C
AC
D016172:DNA Fingerprinting; D018744:DNA, Plant / Q000032:analysis* / Q000302:isolation & purification; D019075:Electrophoresis, Capillary; D005453:Fluorescence; D019105:Random Amplified Polymorphic DNA Technique; D015203:Reproducibility of Results; D019685:Rosales / Q000145:classification / Q000235:genetics*; D012680:Sensitivity and Specificity
D018744:DNA, Plant