Consider this very simple DNA sequence in FASTA format:
>sequence1
nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
ttttttagaaaaattatttttaagaatttttcattttaggaatattgtta
tttcagaaaatagctaaatgtgatttctgtaattttgcctgccaaattcg
tgaaatgcaataaaaatctaatatccctcatcagtgcgatttccgaatca
gtatatttttacgtaatagcttctttgacatcaataagtatttgcctata
tgactttagacttgaaattggctattaatgccaatttcatgatatctagc
cactttagtataattgtttttagtttttggcaaaactattgtctaaacag
If you try converting this to a BLAST database using the 'makeblastdb' command from the latest version of NCBI's BLAST+ suite, you will see the following line included in the output:
Error: (1431.1) FASTA-Reader: Warning: FASTA-Reader: First data line in seq is about 100% ambiguous nucleotides (shouldn't be over 40%)
Now consider what happens if you run the same makeblastdb command on this sequence (which just switches the first two lines of sequence1):
>sequence2
ttttttagaaaaattatttttaagaatttttcattttaggaatattgtta
nnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
tttcagaaaatagctaaatgtgatttctgtaattttgcctgccaaattcg
tgaaatgcaataaaaatctaatatccctcatcagtgcgatttccgaatca
gtatatttttacgtaatagcttctttgacatcaataagtatttgcctata
tgactttagacttgaaattggctattaatgccaatttcatgatatctagc
cactttagtataattgtttttagtttttggcaaaactattgtctaaacag
Although this sequence has the exact same proportion of As, Cs, Gs, Ts, and Ns, it does not produce the error message. What about the following sequence?
>sequence3
nnnac
ttttttagaaaaattatttttaagaatttttcattttaggaatattgtta
tttcagaaaatagctaaatgtgatttctgtaattttgcctgccaaattcg
tgaaatgcaataaaaatctaatatccctcatcagtgcgatttccgaatca
gtatatttttacgtaatagcttctttgacatcaataagtatttgcctata
tgactttagacttgaaattggctattaatgccaatttcatgatatctagc
cactttagtataattgtttttagtttttggcaaaactattgtctaaacag
Well, surprise surprise, this sequence produces the error message again (though it now tells you that the first line 'is about 60% ambiguous nucleotides'). You will still see the same message even if you added 1 billion As, Cs, Gs, and Ts on to the end of sequence 3. This seems to be the code responsible for the error message (taken from this page):