Meanning of flags
- Within genome, one (or more) restriction enzymes may show
overlapped restriction sites.
Example
1
|
|
|
DNA Sequence:
Restriction enzyme:
Recognition sequence:
Cleavage positions:
Cleavage positions recorded:
|
5'-GGGCCC-3'
AspS9I
G'GNC_C
1 and 2
1
|
For a DNA template with
50% C+G content,
GGNCC and GGGCCC
sequences will be present
in the DNA sequence in a
proportion 15:1 |
Example
2
|
|
|
DNA Sequence:
Restriction enzyme:
Recognition sequence:
Cleavage positions:
Cleavage positions recorded:
|
5'-ATTAATTAAT-3'
AseI
AT'TA_AT
2 and 6
2
|
For a DNA template with
50% C+G content,
ATTAAT and
ATTAATTAAT
sequences will be present
in a proportion 255:1. |
When recognition sequences in genome for a specific restriction enzyme
are overlapped, only the first matching recognition sequence (starting
in 5' end) is recorded. As a consecuence, in same cases, the length of
resulting bands may be slightly different from that one shown in our
calculations (bands will be a few bases longer or shorter).
Those errors will not affect bands pattern significantly.
Meanning of flags
- Recognition sequences of restriction enzymes will overlap
sometimes and the resulting bands patterns may be slightly different
from that ones shown in our calculations.
Example
|
|
DNA Sequence:
Restriction enzyme 1:
Restriction enzyme 2:
Cleavage positions:
Cleavage positions recorded:
|
5'-GAATTCCGG-3'
EcoRI (G'AATT_C)
BsiSI (C'CG_G)
1 and 6
6 is not recorded
|
In this case, cleavage by restriction enzyme 2 is not recorded, and it
may change the bands pattern calculated for this combination of
restriction enzymes when the DNA sequence shown in the example is
present in DNA template. As the long sequence (containing recognition
sequences for both restriction enzymes) is likely to happen within
selected genome only a limited number of times, the impact of these
errors in calculations will be limited.
Theoretical calculations have been performed considering digestion of
DNA is sequential: first with restriction enzyme 1 and latter with
restriction enzyme 2. If this procedure is used in the experiment, the
theoretical results are likely to match lab results.
Meanning of flags
- A restriction enzyme cleaves within recognition sequence of the
other restriction enzyme used in the experiment
Example
|
|
DNA Sequence:
Restriction enzyme 1:
Restriction enzyme 2:
|
5'-GAATTC-3'
EcoRI (G'AATT_C)
Sse9I ('AATT_)
|
Theoretical calculations have been
performed considering digestion of DNA is sequential: first with
restriction enzyme 1 and latter with restriction enzyme 2. If this
procedure is used in the experiment, the theoretical results are likely
to match lab results.
