Table 5 |
||||||
|
Pre-restriction of immobilized cDNA |
||||||
|
No pre-restriction |
Partial pre-restriction |
Full pre-restriction |
||||
|
|
||||||
|
Arabidopsis thaliana |
||||||
|
|
||||||
|
Marking enzymes |
FatI |
MboI |
HinPI |
MboI |
FatI |
MboI |
|
|
||||||
|
Releasing enzymes |
MboI |
FatI |
MseI |
MseI |
MboI |
FatI |
|
MseI |
MseI |
TasI |
TasI |
MseI |
MseI |
|
|
TasI |
TasI |
TaqI |
TaqI |
TasI |
TasI |
|
|
|
||||||
|
Sequence coverage |
78 % |
70 % |
52 % |
|||
|
|
||||||
|
Sequence redundancy |
30 % |
30 % |
0 % |
|||
|
|
||||||
|
Average fragment length (nt) |
183 |
237 |
142 |
|||
|
|
||||||
|
PCR reactions |
100 % |
125 % |
36 % |
|||
|
|
||||||
|
Mouse |
||||||
|
|
||||||
|
Marking enzymes |
MboI |
Csp6I |
HinPI |
MboI |
MboI |
Csp6I |
|
|
||||||
|
Releasing enzymes |
Csp6I |
MseI |
TasI |
TasI |
Csp6I |
MseI |
|
MseI |
MboI |
MseI |
MseI |
MseI |
MboI |
|
|
FatI |
FatI |
TaqI |
TaqI |
FatI |
FatI |
|
|
|
||||||
|
Sequence coverage |
87 % |
81 % |
50 % |
|||
|
|
||||||
|
Sequence redundancy |
41 % |
35 % |
0 % |
|||
|
|
||||||
|
Average fragment length (nt) |
270 |
271 |
159 |
|||
|
|
||||||
|
PCR reactions |
100 % |
39 % |
25 % |
|||
|
|
||||||
|
Human |
||||||
|
|
||||||
|
Marking enzymes |
MboI |
Csp6I |
HinPI |
MboI |
MboI |
Csp6I |
|
|
||||||
|
Releasing enzymes |
CspI |
MboI |
MseI |
MseI |
CspI |
MboI |
|
HinPI |
FatI |
TasI |
TasI |
HinPI |
FatI |
|
|
FatI |
HinPI |
TaqI |
TaqI |
FatI |
HinPI |
|
|
|
||||||
|
Sequence coverage |
85 % |
84 % |
63 % |
|||
|
|
||||||
|
Sequence redundancy |
35 % |
40 % |
0 % |
|||
|
|
||||||
|
Average fragment length (nt) |
274 |
292 |
182 |
|||
|
|
||||||
|
PCR reactions |
100 % |
86 % |
61 % |
|||
|
|
||||||
|
Simulation of cDNA-AFLP with sequential digestion protocol on Arabidopsis, human and mouse RefSeq ESTs was performed. Optimal combination of restriction enzymes (2 marking in combination with 3 releasing enzymes) with the highest sequence coverage was used to compare the effect of partial pre-restriction and full pre-restriction options on maximal transcript coverage, sequence redundancy, average fragment length and reduction of PCR effort (relative to the maximal number of PCR by one of the options). |
||||||
|
Weiberg et al. BMC Genomics 2008 9:480 doi:10.1186/1471-2164-9-480 |
||||||
