Figure 1.
Introgression strategies between C. briggsae and C. sp.9.
A. Introgression for the autosome linked marker. B. Introgression for the X chromosome linked marker. See text for more details.
Figure 2.
Flow chart for the selection of C. briggsae specific primers.
See text for more details.
Figure 3.
Intronic structure of abce-1 genes in nematode species.
I: C. sp.9; II: C. elegans; III: C. briggsae; IV: C. brenneri. Exons are denoted as hollow bars with its size in bp labeled inside while introns (not in scale) shown as dashed lines with sizes labeled above.
Figure 4.
Strategy for mapping of a dominant GFP (green bar) marker in C. briggsae.
C. briggsae specific primers (paired arrows) across a chromosome were selected that would specifically amplify the fragment of interest in C. briggsae genome (grey bar) but not that from its homologous region in C. sp.9 (white bar). After multiple generations of backcrossing (introgression) into C. sp.9 background using the GFP positive hybrid progeny, only the C. briggsae genomic fragment that is closely linked to the GFP locus will be retained in the hybrid progeny due to the recombination. The sizes of the GFP linked C. briggsae fragment can be judged by the single worm PCR using a hybrid animal either heterozygous or homozygous (not shown) for the GFP locus as a template. Presence or absence of PCR product will allow simultaneous estimation of the approximate location for the GFP insertion in the C. briggsae genome as well as the calculation of the introgression size in the C. sp.9 background.
Table 1.
Summary of the mapping results.
Figure 5.
A. Test of the primer specificities. A side-by-side comparison of the agarose gel pictures of the PCR products with the genomic DNAs of AF16 (left) or JU1421 (right) as a template. Only a single PCR product was shown for each chromosome with their identities indicated in the bottom. B. Mapping of a GFP locus onto the middle of chromosome II using the GFP positive hybrid progeny (after 15 generations of introgression) as a template. A total of 14 pairs of primers (ordered from left to right based on their genomic coordinates) were used and only a single pair of primers gave rise to PCR product with expected size, indicating the GFP insertion site is located between the boundaries defined by its two adjacent pairs of primers. Chromosomal arms are defined into “Left”, “Middle” and “Right” based on their relative genomic coordinates.
Figure 6.
Amplification of the PCR products with the sizes expected for AF16 using the genomic DNAs of EG5268 but not that of JU1421 as a template for two out the 7 pairs of primers.
A side-by-side comparison of PCR amplification using the genomic DNAs of JU1421 (left of each pair) or EG5268 (right of each pair) with the primers specific for the chromosome I. Two unexpected amplifications with EG5268 were shown. Only results of seven pairs of primers (A to G) were shown. All of the primers gave rise to the expected PCR amplifications with AF16 genomic DNA as a template (data not shown).
Figure 7.
PCR amplification of discontinuous genomic fragments on the same chromosome using animals from one introgression lines as a template.
Shown were the PCR amplifications with six pairs of primers (A–F) specific for the chromosome X using a GFP positive adult worm from two different introgression lines as a template after 15 generation of introgression. The PCR product was ordered by the genomic coordinates of the selected primers from the left to the right arm. Note the amplifications of the discontinuous regions as indicated by the primers D and F but not by E (panel I) as opposed to the amplifications of a continuous region as indicated by the primers E and F but not the primer D (panel II).