ArabidopsisRILs

Existing RIL sets

We have been using the Cvi/Ler, Nd/Col, Kas/Col, and Bay/Shad RIL sets. Many others are currently available from the stock center.  We have mapped QTL affecting hypocotyl length and/or flowering time.

New RIL sets

We are also generating several sets of Recombinant Inbred Lines (RILs) by crossing different accessions to the Columbia (Col) reference wild-type strain. See our letter regarding our policy about distribution of these RILs.

Progress as of Jan21st, 2008

* Genotyping is being used to determine the exact parentage of the cross.

We now have > 100 markers for Est/Col RILs.

Protocol:

To increase the number of recombination events in each set, we have performed for most sets several generations of random intercrossing before beginning with single-seed descent (see table). Briefly, beginning with the F2, 75 (or 150 VanC) random, non-overlapping pairs of individuals were selected for crosses. Two plants from each cross were kept for the next round of random intercrossing.

To generate recombinant inbred lines, each set of F2s or advanced intercross lines (AILs) is being taken through six additional generations of random single-seed descent, which should yield lines that are more than 98% homozygous across the genome. For lines with relatively uniform flowering times (not requiring vernalization), 20 seeds from each plant are imbibed in water for 4 days at 4°C. After stratification, seeds are placed on soil in 12-pot flats, with 4 lines per pot (one line in each corner of the pot). For each line, only the seedling closest to the corner of the pot is kept, thereby randomly propagating seedlings with respect to growth characteristics to avoid selection bias. For lines requiring vernalization, 20 seeds per line are sterilized, soaked overnight in water, and then plated on 1/2 MS phytagar plates; plates with gridlines are being used, and one line is plated per gridded square. Seeds are then stratified for an additional 4 days in the dark at 4°C, moved to 23°C light for 3 days to induce germination, and placed at 4°C in a short-day incubator for 4 to 6 weeks. After vernalization, one seedling per line is chosen (the upper-leftmost seedling to prevent selection bias) and transplanted to soil.

After the sixth round of selfing, the RILs will be genome-wide genotyped. When the lines are finished, we will deposit them in the stock center. At the present time, we cannot give a firm estimate when this will happen, but likely sometime during 2004 for the first set of lines, if not sooner. We will provide an update at this site regarding submission status of the lines on this site.