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HYDEN v1.0 (Windows)
Copyright C. Linhart, R. Shamir and RAMOT (Tel Aviv University), 2002.
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Dna file (fasta format) ............................................ ..\HGP_50genes.fasta
Maximal number of primer pairs to design ........................... 2
Length of 5' primer ................................................ 25
Length of 3' primer ................................................ 25
Maximal degeneracy of 5' primer .................................... 5000
Maximal degeneracy of 3' primer .................................... 30000
Sequence positions for 5' primer ................................... 0 to 300
Sequence positions for 3' primer ................................... -1 to -350
Maximal number of mismatches in 5' end ............................. 2
Maximal number of mismatches in 3' end ............................. 2
Maximal number of mismatches in both ends combined ................. 3
Number of DNA sequences for entropy estimation ..................... 30
Number of base primers to run contraction/expansion algorithms ..... 500
Number of best candidates to run greedy improvement algorithm ...... 50
Read 50 DNA sequences from file ..\HGP_50genes.fasta.
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Designing a primer pair for 50 DNA sequences.
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==> Designing 5' primer between positions 0 and 300.
Phase 1: Finding conserved base primers (blocks with low entropy score).
Computing entropy of base primers according to 30 sequences.
Lowest entropy (on 30 sequences) received at position 166 in sequence #4: 15.682
Highest entropy (on 30 sequences) received at position 244 in sequence #47: 42.342
Computing entropy of best 2500 base primers according to all 50 sequences.
Lowest entropy (on all 50 sequences) received at position 160 in sequence #43: 16.703
Highest entropy (on all 50 sequences) received at position 16 in sequence #2: 35.825
Phase 2: Running contraction/expansion algorithms on best 500 base primers.
Got 508 primers from contraction/expansion algorithms.
Primers cover between 30 and 45 sequences (with up to 2 mismatches).
Phase 3: Running greedy improvement algorithm on best 50 primers.
Best 5' primer covers 46 sequences (out of 50) with up to 2 mismatches:
Primer: length 25, degeneracy 4608
CTNSAYDCNCCYATGTAYTTYTTHC
A + + + + + + +
C+ ++ + +++++ + + ++
G ++ + + +
T ++ ++ + + + + +++++++
==> Designing 3' primer between positions -1 and -350;
working only on 46 sequences that are matched by the 5' primer.
Phase 1: Finding conserved base primers (blocks with low entropy score).
Computing entropy of base primers according to 30 sequences.
Lowest entropy (on 30 sequences) received at position 853 in sequence #39: 17.337
Highest entropy (on 30 sequences) received at position 592 in sequence #33: 42.626
Computing entropy of best 2500 base primers according to all 46 sequences.
Lowest entropy (on all 46 sequences) received at position 855 in sequence #17: 18.270
Highest entropy (on all 46 sequences) received at position 818 in sequence #20: 32.000
Phase 2: Running contraction/expansion algorithms on best 500 base primers.
Got 401 primers from contraction/expansion algorithms.
Primers cover between 30 and 41 sequences (with up to 2 mismatches).
Phase 3: Running greedy improvement algorithm on best 50 primers.
Best 3' primer covers 43 sequences (out of 46) with up to 2 mismatches (printed inverted):
Primer: length 25, degeneracy 27648
TCCTBADRSTRTARATNANNGGDTT
A +++ + +++ ++++ +
C ++ + + + ++
G + +++ + + + +++++
T+ ++ + + + ++ ++ +++
Best 3' primer covers a total of 46 sequences (out of 50) with up to 2 mismatches.
==> Primer pair #1 matches 34 DNA sequences (that are not matched by previous primers)
with up to 3 mismatches (in both sides combined).
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Designing a primer pair for 16 remaining DNA sequences.
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==> Designing 5' primer between positions 0 and 300.
Phase 1: Finding conserved base primers (blocks with low entropy score).
Computing entropy of base primers according to 16 sequences.
Lowest entropy (on 16 sequences) received at position 168 in sequence #0: 22.746
Highest entropy (on 16 sequences) received at position 94 in sequence #3: 39.822
Computing entropy of best 2090 base primers according to all 16 sequences.
Lowest entropy (on all 16 sequences) received at position 168 in sequence #0: 22.746
Highest entropy (on all 16 sequences) received at position 94 in sequence #3: 39.822
Phase 2: Running contraction/expansion algorithms on best 500 base primers.
Got 755 primers from contraction/expansion algorithms.
Primers cover between 1 and 12 sequences (with up to 2 mismatches).
Phase 3: Running greedy improvement algorithm on best 50 primers.
Best 5' primer covers 12 sequences (out of 16) with up to 2 mismatches:
Primer: length 25, degeneracy 4608
MCCCCATGTAYTTYTTYCYBDSMAN
A+ + + + +++
C+++++ + + ++++ ++ +
G + +++ +
T + + +++++++ +++ +
==> Designing 3' primer between positions -1 and -350;
working only on 12 sequences that are matched by the 5' primer.
Phase 1: Finding conserved base primers (blocks with low entropy score).
Computing entropy of base primers according to 12 sequences.
Lowest entropy (on 12 sequences) received at position 882 in sequence #10: 22.600
Highest entropy (on 12 sequences) received at position 669 in sequence #3: 37.813
Computing entropy of best 2313 base primers according to all 12 sequences.
Lowest entropy (on all 12 sequences) received at position 882 in sequence #10: 22.600
Highest entropy (on all 12 sequences) received at position 669 in sequence #3: 37.813
Phase 2: Running contraction/expansion algorithms on best 500 base primers.
Got 807 primers from contraction/expansion algorithms.
Primers cover between 1 and 9 sequences (with up to 2 mismatches).
Phase 3: Running greedy improvement algorithm on best 50 primers.
Best 3' primer covers 10 sequences (out of 12) with up to 2 mismatches (printed inverted):
Primer: length 25, degeneracy 20736
ATKRCWGHYTTBAMHWCHTYATTYC
A+ + + + ++++ + +
C + ++ + ++ ++ + ++
G ++ + +
T ++ + +++++ ++ +++ +++
Best 3' primer covers a total of 10 sequences (out of 16) with up to 2 mismatches.
==> Primer pair #2 matches 6 DNA sequences (that are not matched by previous primers)
with up to 3 mismatches (in both sides combined).
Found 2 pairs of primers:
(coverage of each primer pair is with respect to the
DNA sequences that are not covered by the previous primers;
the last column shows the total accumulated coverage)
Pair | 5' primer | 3' primer (inverted) | coverage | % (acc.)
------+---------------------------+---------------------------+----------+----------
0 | CTNSAYDCNCCYATGTAYTTYTTHC | TCCTBADRSTRTARATNANNGGDTT | 34 | 68 %
1 | MCCCCATGTAYTTYTTYCYBDSMAN | ATKRCWGHYTTBAMHWCHTYATTYC | 6 | 80 %
------+---------------------------+---------------------------+----------+----------