ABSTRACT
Vectors ....................................................................................................... 529 30.3.1.3 Preparation of Blunt-End DNA .................................................................. 531 30.3.1.4 Preparation of Insert DNA Lacking the 5′ Phosphate Group .................... 531
30.3.2 Blunt-End Ligation of Plasmid Vectors and Inserts of Antisense and Sense Orientation ............................................................................................... 533 30.3.2.1 Reagents Needed ........................................................................................ 534
30.3.3 Transformation of Appropriate Strain of Bacteria to Amplify the Recombinant Plasmids .................................................................................................................... 534
30.3.4 Selection of Plasmids with Antisense and Sense Orientations ................................. 534 30.3.5 Gene Transfer and Expression of Antisense RNA ................................................... 535
30.4 New Potential of Double-Stranded RNA for the Suppression of Gene Expression ............. 536 30.4.1 Insights into the Mechanisms of Double-Stranded RNA ......................................... 536 30.4.2 RNAi via Feeding in C. elegans ............................................................................... 536
30.4.2.1 Equipment and Reagents Needed .............................................................. 537 30.4.3 RNAi via Microinjection in C. elegans .................................................................... 538
30.4.3.1 Equipment and Reagents Needed .............................................................. 539 30.4.4 RNAi via Soaking in C. elegans ..............................................................................540 30.4.5 Inducible or Tissue-Speci c RNAi in C. elegans ....................................................540
30.5 Overview of Mammalian RNAi Pathway and RNAi Technologies in Mammalian Cell Cultures ......................................................................................................................... 541 30.5.1 Mammalian RNAi .................................................................................................... 541 30.5.2 Overview of Mammalian Cell Culture Transfection Methods ................................. 542
