PROBING RIBOSOMAL FUNCTION

化学合成的DNA低聚物长度小于10个碱基

Chemically-synthesized DNA oligomers less than ten bases long are to be used to probe the functions of various regions of ribosomal RNA in situ. The DNA oligomers will be synthesized so as to be complementary to certain exposed regions of rRNA, hybridized to those regions, assayed for specific binding and then the complex will be assayed to ascertain if the function of the ribosome has been impaired by the presence of the DNA probe. Specific binding of the complementary DNA to a specific rRNA site will be assured by using RNase H to clip the rRNA in the heteroduplex region and then sequencing the resulting fragments for short distances in either direction from the slip. The functional assays to be used will include protein synthesis, tRNA binding to both the P site and the A site, initiation complex formation and mRNA binding assays. It is expected that discrete functions of specific regions of rRNA in the intact ribosome will be determined in this manner.