NOVEL SCREEN FOR RESTRICTION ENDONUCLEASES

限制性内切酶是基因工程的关键试剂。 虽然已经有200多种特异性， 需要更多识别新序列。比如说， 另外的识别八核苷酸序列的酶将被 许多疾病基因含有突变， 如果使用适当的限制性内切酶， available.目前的生化筛选方法效率低下， 费力而且很少成功。这些检测发现 在四种微生物中只有一种被检测到， 发现新的特异性急剧下降（只有3个在 去年）。目前的DNA测序工作表明， 细菌至少有一个，可能更多，限制- 修改（RM）系统。该提案提供了一个新的， 基于计算的方法，筛选每日更新的 潜在限制酶基因的基因库，并有助于 确定那些可能具有新特异性的。候选人将被 进行生化筛选以检验预测结果。新特异性 将被充分表征并提供销售。 建议的商业应用这项研究将导致 发现具有新的限制性内切酶 特殊性 它还将确定一些基因的克隆和序列， 以前未克隆的酶。 新的酶将出售给 生物技术产业和学术研究界。

Restriction endonucleases are key reagents for genetic engineering. Although more than two hundred specificities already exist, there is a need for more that recognize novel sequences. For instance, additional enzymes recognizing octanucleotide sequences would be useful for mapping and many disease genes contain mutations that could easily be detected if an appropriate restriction enzyme were available. Current biochemical screening methOds are inefficient, laborious and rarely successful. These assays find restriction enzymes in only one of four microorganisms examined and the rate of finding new specificities has dropped dramatically (only 3 in the last year). Current DNA sequencing efforts suggest that all bacteria possess at least one, and probably many more, Restriction- Modification (RM) systems. This proposal offers a new, computationally-based approach that screens the daily updates of GenBank for potential restriction enzyme genes and helps to identify those that may have new specificities. Candidates will be screened biochemically to test the predictions. New specificities will be fully characterized and offered for sale. PROPOSED COMMERCIAL APPLICATIONS This research will lead to the discovery of new restriction endonucleases with novel specificities. It will also identify clones and sequences for some previously uncloned enzymes. The new enzymes will be sold both to the Biotechnology industry and to the academic research community.