Direct tissue blotting for detection of rice viruses
1994
Miranda, G.J. | Koganezawa, H.
Most serological diagnostic methods for plant viruses require attraction of plant sap. Such a process limits its use in a field. Direct tissue blotting which does not need a sap extraction step, was applied for field testing of rice viruses such as rice tungro bacilliform virus (RTBV), rice tungro spherical virus (RTSV), rice grassy stunt virus (RGSV), and rice ragged stunt virus (RRSV). Rice plant parts, culm, sheath, and leaf, were cut by a razor blade. The cut surface was directly blotted by pressing it gently onto a 0.45 micromole pore size nitrocellulose membrane. Signal for virus infection was detected using direct and indirect immunoblotting. Of the three tissues, the sheath samples gave the best signal. The indirect method is more sensitive than the direct one in detecting all viruses. Optimum dilutions of immunoglobulin (1 g G) for the indirect method were 1/4000 for RGSV and 1/1000 for RTBV, RTSV, and RRSV. After adding the substrates, initial reaction started within 15-30 min for RGSV and 30-45 min for RTBV, RTSV, and RRSV. The whole procedure took 4-5 h. The results showed that tissue blotting is specific, sensitive, rapid, and is a convenient method to assay a large number of samples
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