[理学]基因工程原理Chapter 2.ppt

Chapter 2 Gene manipulation: an all-embracing technique 2-1 Introduction 1. Some developments enable science to lead forward and increase the potential for innovation 2. Gene manipulation Sophisticated in vivo genetics Applied to Sophisticated in vitro techniques New combinations of heritable material are constructed by the insertion of nucleic acid molecules, though virus, bacterial plasmid or other vector system outside cell, then the incorporated vector enters host organism and can multiply inside the host, showing unnatural. Simply speaking, gene manipulation permits stretches of DNA to be isolated (cloned) from their host organism and propagated in the same or a different host. 2-2 Sequencing Steps: cloning sequencing scanning deducing protein sequence based on nucleotide sequence . Scanning If a fragment sequenced, then put the sequence into a searching engine on the internet: BLAST (/), for searching its homologue and possible function. BLAST (Basic Local Alignment Search Tool,局域相似性比对分析工具) How to BLAST? 1. Nucleotide-nucleotide BLAST, blastn); 2. Nucleotide query-protein, blastx But remember: FASTA format is required. Result for the search Gene prediction 1. ORF finder is usually used for this purpose at the beginning; 2. Function domain prediction. 3. Many bio-software are convenient on line or available from some companies: Bio-edit, DNAman, etc. If you have a specific fragment or gene, you will know its digestion site, start and stop codons, inverted palindromes, sequence repeats, Z-DNA and so on. Promoter prediction / is available on the internet; The 2kb sequence before the initiation codon of an ORF should be used for the prediction. Deducing protein sequence By software! It is so easy job! What you do is just　to “Click” some buttons in your computer. Motif: continuous sequence of amino acids or DNA so as to form a fragment. It is very difficult to predict a protein from three dimensional structure f