Fig 4. ZIPT-7.1 is expressed and functions in the germ line to control sperm activation. (A) Bands represent transcript levels determined bysemiquantitative RT-PCR. Genotypes are labeled above, target transcripts at the left, and germ line phenotypes below;
act-1 is a loading control. Foreach lane, RNA was isolated from batches of five worms (see Materials and methods). The alleles were
him-5(
e1490),
fem-3(
q96),
fem-1(
hc17ts),
fog-1(
q253ts), and
glp-4(
bn2ts). (B) Diagram illustrating the insertion site of the GFP coding sequence into the endogenous
zipt-7.1 locus to generate thezipt-7.1
(ibp18)strain. (C) Sets of photomicrographs from two extruded male gonads. The location of the images in the gonad arm is indicated with ared box on the cartoon; distal is up and proximal to the right. The bright-field images show cell morphology (left), the antibody stain shows theexpression of GFP::ZIPT-7.1 (middle), and the DAPI stain shows cell nuclei (right). Genotypes were
him-5(
e1490) and
him-5(
e1490)
zipt-7.1(
ibp18).Scale bar is 10 um. (D) Reproductive defects caused by
zipt-7.1 RNAi treatment. The F1 progeny of mothers injected with dsRNA were each scoredfor whether they laid eggs, oocytes, or both. Dotted lines show the average percentage for each phenotype and sum to 100%, the size of the boxesindicates 95% confidence limits, and the color indicates the phenotype according to the key. The
rrf-1 genotype and RNAi treatment are indicated atthe bottom (N = 200, 248, and 315, left to right). Wild-type animals are susceptible to RNAi in germ cells and most somatic cells, whereas the
rrf-1mutant animals are susceptible to RNAi in germ cells but resistant in most of the somatic cells [17-19], as indicated in the diagram. The individualnumerical values for panel D can be found in S1 Data. DIC, differential interference contrast; dsRNA, double-stranded RNA; GFP, green fluorescentprotein; Oo, oocyte; Sp, sperm; RNAi, RNA interference; RT-PCR, Reverse transcription polymerase chain reaction.