18S Ribosomal RNA Detection on Northern Blot Employing a Specific Oligonucleotide

mation of yeast (Table 1B). An increased number, 43, of the transformants were recombinants. When the DNA linkers were omitted, fewer transformants and no recombinants were obtained (Table 1C). In further cloning experiments using other genes (not shown), we have found that the indirect method of detection of recombinants, by loss of TRP1, is a useful method to help identify yeast transformants that contain exclusively recombinant plasmids. In each case, the DNA sequence of clones was confirmed directly. For cloning of DNA fragments that did not have a phenotype such as fluorescence, this greatly reduced the labor required to obtain recombinant plasmids. This method is particularly applicable when the end product is a yeast expression vector because it eliminates conventional cloning steps in bacteria, minimizes PCR amplification errors in the cloned DNA, and does not require unique restriction sites in the donor plasmid substrate.