Kwik fil 1b100f 4

To obtain germ-line transformation of C. elegans expression plasmids with a selection marker were injected in the cytoplasm of the syncytial gonad. Microinjection was done generally by the method described in WormBook (Evans 2006 ). DNA solution was prepared by mixing the expression plasmid (10-20 μg/ml) and pRF4 (rol-6(su1006)) (100 μg/ml). Final concentration of DNA mixture was adjusted to 200 μg/ml with pBluescript. The DNA solution was loaded to a needle made from a borosilicate glass capillary (Kwik-Fil™ (1B100F-4), World Precision Instruments, Inc., Sarasota, FL) using microloader tip (Eppendorf AG, Germany). The needle was made using Sutter micropippette Puller (P-2000, Sutter Instrument, Novato, CA). DNA solution was injected to 20-30 well-fed gravid hermaphrodites using Axio Observer.A1 inverted microscope (Carl Zeiss Microscopy GmbH, Germany) equipped with three-axis hanging joystick oil hydraulic micromanipulator (MMO-202ND, Narishige, Japan), and microINJECTOR™ (Tritech Research, Inc., Los Angeles, CA).