Rubber Tree (Hevea brasiliensis Muell. Arg)

Rubber tree (
Hevea brasiliensis
Muell. Arg.) is an important industrial crop for natural rubber production. At present, more than 9.5 million hectares in about 40 countries are devoted to rubber tree cultivation with a production about 6.5 million tons of dry rubber each year. The world supply of natural rubber is barely keeping up with a global demand for 12 million tons of natural rubber in 2020. Tapping panel dryness (TPD) is a complex physiological syndrome widely found in rubber tree plantations, which causes severe yield and crop losses in natural rubber producing countries. Currently, there is no effective prevention or treatment for this serious malady. As it is a perennial tree crop, the integration of specific desired traits through conventional breeding is both time-consuming and labour-intensive. Genetic transformation with conventional breeding is certainly a more promising tool for incorporation of agronomically important genes that could improve existing
Hevea
genotype. This chapter provides an
Agrobacterium
-mediated transformation protocol for rubber tree using immature antherderived calli as initial explants. We have applied this protocol to generate genetically engineered plants from a high yielding Indian clone RRII 105 of
Hevea brasiliensis
(Hb). Calli were co-cultured with
Agrobacterium tumefaciens
harboring a plasmid vector containing the Hb superoxide dismutase (SOD) gene and the reporter gene used was β-glucuronidase (GUS) gene (
uidA
). The selectable marker gene used was neomycin phosphotransferase (
npt
II) and kanamycin was used as selection agent. We found that a suitable transformation protocol for
Hevea
consists of a 3-d co-cultivation with
Agrobacterium
in the presence of 20 m
M
acetosyringone, 15 m
M
betaine HCl, and 11.55 m
M
proline followed by selection on medium containing 300 mg/L kanamycin. Transformed calli surviving on medium containing 300 mg/L kanamycin showed a strong GUS-positive reaction. Upon subsequent subculture into fresh media, we obtained somatic embryogenesis and germinated plantlets, which were found to be GUS positive. The integration of
uidA
,
npt
II, and HbSOD transgenes into
Hevea
genome was confirmed by polymerase chain reaction (
PCR
) as well as Southern blot analysis.