INTERPRETATION OF HYBRIDISATION RESULTS
The filters are labelled at the top left hand corner (LA13-1and LA13-2).
This label serves as an orientation mark. The enclosed sheets [i,
and tables (ii) and (iii)] provide detailed information about the
layout of clones on each filter.
There is one large filter and one small filter supplied. 12 plates have
been gridded onto the small filter which is a third of the size of the
large filter.
Sheet (i) shows a typical 22 x 22 cm membrane with 36,864 spotted clones.
Each filter is divided into 6 panels (indicated by 1, 2 ......6
). The panels are numbered from left to right and eight 384-well plates
are gridded, in duplicate, in each panel. Within each panel, numbers 1-24
correspond to the 24 columns and letters A-P are the 16 rows of the eight
384-well plates in the panel. Each dot represents a clone from a single
microtitre plate well. A group of 16 dots corresponds to a 4x4 array of
duplicate clones from the 8 microtitre plates (8 x 2). Thus each panel
consists of 6144 clones (384 x 8 x 2). Table (ii) shows the order of the
8 plates (in duplicate) within each panel on the filter. Panel 1
contains plates 1-8, panel 2 contains 9-16 and so on. This plate
order has been maintained on all the membranes, e.g.
|
Membrane LA13-1
|
Panel 1
|
Membrane LA13-1
|
Panel 2
|
|
|
plates
|
3
|
5
|
6
|
8
|
|
plates
|
11
|
13
|
14
|
16
|
|
|
|
2
|
7
|
3
|
4
|
|
|
10
|
15
|
11
|
12
|
|
|
|
6
|
4
|
8
|
5
|
|
|
14
|
12
|
16
|
13
|
|
|
|
2
|
1
|
7
|
1
|
|
|
10
|
9
|
15
|
9
|
....etc.
The interpretation of positive signal(s) can be done by following the
procedure given below:
- Identify the membrane on which the positive signal is located (in
duplicate).
- Use the label position to orientate the filter and identify the panel
in which the positive clones are located.
- Work out the microtitre co-ordinates of the 4 x 4 array by referring
to the row (A-P) and column (1-24) designations.
- Within the identified 4 x 4 array determine which plate number contains
the positive clone.
Check that any two signals fall in one of the following patterns:
Use either table (ii) or table (iii) to identify which plate number
is giving the positive signal.
- Record the coordinates of putative positives by following the convention
PLATE - ROW and COLUMN (e.g. 16-L15). When requesting this clone using
our WWW request form the clone ID must be preceded by the short library
name (e.g. LA1316L15) and clearly identified as the result of Geneservice
filter screening.
Version 1.0 10th June 1999
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