American Chestnut [Castanea dentata (Marsh.) Borkh].
The key to successful transformation of American chestnut is having the correct combination of explant tissue, selectable and scorable markers, and a reliable regeneration system. Rapidly dividing somatic embryos, growing as proembryogenic masses, are a suitable tissue; the bar gene is a suitable selectable marker in conjunction with 1.0 to 10 mg/L phosphirothricin (PPT); and the mgfp5-ER gene is an effective nondestructive scorable marker. We have also found that the more gently the somatic embryos are treated during the inoculation and co-cultivation steps, the higher the transformation efficiency. The average transformation efficiency that can be expected using the described protocol is approx 20 stable and embryogenic transformation events/g of somatic embryo tissue. Cell line and batch-to-batch deviations both upward and downward should be expected. Finally, somatic embryos can be induced to form shoots, which can then be micropropagated and acclimatized.
Maynard
Charles A
CA
SUNY College of Environmental Science and Forestry, State University of New York, Syracuse, NY, USA.
Polin
Linda D
LD
LaPierre
Sharon L
SL
Rothrock
Ronald E
RE
Powell
William A
WA
D000064:Acclimatization; D018929:Cell Culture Techniques; D018920:Coculture Techniques; D003470:Culture Media; D029962:Fagaceae / Q000033:anatomy & histology / Q000196:embryology / Q000235:genetics*; D005819:Genetic Markers; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000032:analysis; D018517:Plant Roots / Q000235:genetics / Q000254:growth & development; D018520:Plant Shoots / Q000235:genetics / Q000254:growth & development; D012231:Rhizobium / Q000166:cytology / Q000235:genetics*; D012639:Seeds / Q000235:genetics / Q000254:growth & development; D046509:Tissue Culture Techniques; D014170:Transformation, Genetic
D003470:Culture Media; D005819:Genetic Markers; D049452:Green Fluorescent Proteins