High-density filter layout

Each 22x22cm filter contains 18,432 independent clones spotted in duplicate in a 4x4 array pattern for a total of 36,864 colonies on each filter. Three to four anchor spots are present on each filter (see detail page for each species to determine location).

Each filter is organized into six separate fields (Figure 1). Each field is divided into 384 well positions (the same layout as a 384-well plate), having 24 columns (1-24) and 16 rows (A-P), (Figure 2). Each field represents clones from eight 384 well plates (see filter detail page) spotted in duplicate (Figure 1B).

The A1 corner is marked by a number in the top right hand corner on the back of each filter and should be used to orient the filter correctly.

Determining the address of a positive clone

Once you have identified a positive clone, you need to determine its plate number and well position.

1. Holding the film so the A1 corner is on the top right, determine which field the positive clone is located in (for details on the particular field layout please refer to the appropriate filter detail sheet for each library).

2. Orient the transparent overlay perpendicular to the field and determine the well position of your positive clone.

3. Use the appropriate vector sheet to determine the spotting pattern of each positive clone. Once the spotting pattern has been determined, use the appropriate filter detail sheet to locate which plate the positive clone originated from.

A worked example

1. This example uses the filter detail sheet for tilapia library #3 (see below).

2. A positive clone is found in field #2.

3. The positive clone was spotted following pattern number 8.

4. As seen in Figure 1, using the transparent overlay sheet as a guide, the positive clone is located in well H6.

5. Using the filter detail sheet, determine the plate number: pattern #8, in field 2, corresponds to plate number 92.