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in the Yellow Sea population, respectively. The
Z
-test showed that
d
N
was significantly
greater than
d
S
in the PBR (
d
N
/d
S
= 5.8,
P
= 0.002), but could not reach the significant level in
the non-PBR (
d
N
/d
S
=1.6,
P
= 0.188) (Table 2).
Allelic Diversity of the Baiji at Three MHC Genes
DRA
Three
DRA
unique sequences were identified from 75 clones of 15 baiji (Figure 1a). All
individuals have no more than two sequences, which suggested that only one
DRA
locus was
amplified in each sample. Sequences are available in GenBank under Accession Nos.
DQ851844-DQ851846. The allele
Live-DRA*03
most likely represented a pseudogene, as
evidenced by the absence of a base in the middle part of the sequence. This allele, therefore,
was not included in the following analyses. The allele frequency of
Live-DRA*01
and
Live-
DRA*02
was 0.633 and 0.367, respectively. In the 189 bp
DRA
sequence, only two nucleotide
sites (1.1%) were variable in the baiji. An alignment of the 63 unique inferred amino acid
sequences revealed two variable sites (3.2%), both of which represented nonsynonymous
substitutions in the non-PBR (Figure 1a & Table 2).
DQB
Eight
DQB
alleles of
L. vexillifer
were identified (Figure 1b). Sequences are available in
GenBank under Accession Nos.:
Live-DQB*4
: AY177153;
Live-DQB*5
: AY177283;
Live-
DQB*8
: AY177286;
Live-DQB*11
: AY177289;
Live-DQB*13
: AY177291;
Live-DQB*16
:
AY333383;
Live-DQB*28
: AY333395;
Live-DQB*29
: AY333396). The variability was 16 of
172 (9.30%) in nucleotide sequences and 9 of 57 (15.79%) in amino acid residues (Figure
1b). The number of pairwise nucleotide differences between pairs of sequences ranged from 1
(
Live-DQB
*
8
vs.
Live-DQB
*
11
) to 16 (
Live-DQB*28
vs.
Live-DQB
*
11
), and the number for
amino acid varied from 0 (
Live-DQB
*
8
vs.
Live-DQB
*
11
) to 9 (
Live-DQB*28
vs.
Live-
DQB
*
11
). Considerable sequence variation was also evident because the rate of
nonsynonymous substitutions was more than eight times higher than that of synonymous
substitutions (
P
= 0.033,
Z
-test of positive selection) in the putative PBR, while the rate
decreased to 0.2 (
P
= 0.136,
Z
-test of positive selection) in the Non-PBR (Table 2).
MHC-I
A 147 bp fragment of the MHC-I exon 2 was amplified from each of 15 baiji. A total of
111 clones were sequenced and six unique sequences,
Live-I*01
to
Live-I*06
(GenBank
Accession Nos. DQ851847-DQ851852), were identified (Figure 1c). We named these
sequences as MHC alleles although we were aware that they stemmed from at least three
different loci rather than a single specific locus. Ten out of 15 individuals had three to five
discrepant sequences. But in further analyses, we considered all sequences as if they would be
alleles of one locus. None of the sequences contained insertions/deletions (or indels) or stop
codons, suggesting that all sequences come from functional alleles in the genome. Thirty-five
out of 147 (23.8%) nucleotide positions were variable. The number of pairwise nucleotide
differences between pairs of alleles ranged from six (
Live-I*04
vs.
Live-I*06
) to 30 (
Live-
I*05
vs.
Live-I*06
). In the deduced amino acid sequences, 21 out of 49 (42.9%) were variable
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