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Isolation and Characterization of the Etheostoma tallapoosae (Teleostei: Percidae) CENP-A Gene.

Fountain DM, Kral LG - Genes (Basel) (2011)

Bottom Line: As a result of this study we have demonstrated that the Tallapoosa darter CENP-A gene HFD sequences can be isolated from genomic DNA by nested PCR in a manner that does not lead to the amplification of the highly sequence related histone H3 gene.These adjacent gene sequences provide us with a primer binding sites for PCR isolation of the CENP-A gene from other percid species of fishes.An initial comparison of three percid species shows that the N-terminal tail of the percid CENP-A gene shows adaptive evolution.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of West Georgia, Carrollton, GA 30118, USA. dfountain82@gmail.com.

ABSTRACT
Both centromeric alpha-satellite sequences as well as centromeric protein A (CENP-A) are highly variable in eukaryotes. CENP-A, a histone H3 variant, is thought to act as the epigenetic "mark" for assembly of centromeric proteins. While most of the histone fold domain (HFD) of the CENP-A is fairly well conserved, a portion of this HFD as well as the N-terminal tail show adaptive variation in both plants and animals. Such variation may establish reproductive barriers that may lead to speciation. The family Percidae contains over 200 species most of which are within the subfamily Etheostomatinae. This subfamily represents a species rich radiation of freshwater fishes in North America and these species exhibit both allopatric and sympatric distributions. In order to study the evolution of CENP-A in percid fish species, we have isolated and characterized the CENP-A gene from Etheostoma tallapoosae by PCR based gene walking. As a result of this study we have demonstrated that the Tallapoosa darter CENP-A gene HFD sequences can be isolated from genomic DNA by nested PCR in a manner that does not lead to the amplification of the highly sequence related histone H3 gene. We also demonstrated that PCR based walking can be subsequently used to isolate the rest of the CENP-A gene and adjacent gene sequences. These adjacent gene sequences provide us with a primer binding sites for PCR isolation of the CENP-A gene from other percid species of fishes. An initial comparison of three percid species shows that the N-terminal tail of the percid CENP-A gene shows adaptive evolution.

No MeSH data available.


Related in: MedlinePlus

Comparative sequence analysis of CENP-A of P. austroperka (Pa), P. roanoka (Pr), and E. tallapoosae (Et). The red underline indicates a region within the N-terminal tail where Ka > Ks.
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f4-genes-02-00829: Comparative sequence analysis of CENP-A of P. austroperka (Pa), P. roanoka (Pr), and E. tallapoosae (Et). The red underline indicates a region within the N-terminal tail where Ka > Ks.

Mentions: An alignment of the coding sequences from E. tallapoosae, P. austroperka, and P. roanoka is shown in Figure 4. To determine if evidence of positive selection could be obtained from this sequence comparison, pairs of these sequences were initially subject to sliding window analysis utilizing the SWAKK web server [18]. This initial analysis showed that a KA > KS signal was obtained in a 20 amino acid segment in the middle of the N-terminal tail (underlined in red in Figure 4). Statistical significance of this positive selection signal was tested by both the Z-test of Selection and by Fisher's Exact Test of Selection [19] where the numbers of synonymous and non-synonymous differences between sequences were estimated using the Nei-Gojobori method [20]. As indicated in Table 4 and Table 5, both of the tests show that in this portion of CENP-A the rate of non-synonymous substitutions is significantly greater than the rate of non-synonymous substitutions between E. tallapoosae and the two Percina species (p < 0.05). The remainder of the sequence is under purifying selection (data not shown).


Isolation and Characterization of the Etheostoma tallapoosae (Teleostei: Percidae) CENP-A Gene.

Fountain DM, Kral LG - Genes (Basel) (2011)

Comparative sequence analysis of CENP-A of P. austroperka (Pa), P. roanoka (Pr), and E. tallapoosae (Et). The red underline indicates a region within the N-terminal tail where Ka > Ks.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3927593&req=5

f4-genes-02-00829: Comparative sequence analysis of CENP-A of P. austroperka (Pa), P. roanoka (Pr), and E. tallapoosae (Et). The red underline indicates a region within the N-terminal tail where Ka > Ks.
Mentions: An alignment of the coding sequences from E. tallapoosae, P. austroperka, and P. roanoka is shown in Figure 4. To determine if evidence of positive selection could be obtained from this sequence comparison, pairs of these sequences were initially subject to sliding window analysis utilizing the SWAKK web server [18]. This initial analysis showed that a KA > KS signal was obtained in a 20 amino acid segment in the middle of the N-terminal tail (underlined in red in Figure 4). Statistical significance of this positive selection signal was tested by both the Z-test of Selection and by Fisher's Exact Test of Selection [19] where the numbers of synonymous and non-synonymous differences between sequences were estimated using the Nei-Gojobori method [20]. As indicated in Table 4 and Table 5, both of the tests show that in this portion of CENP-A the rate of non-synonymous substitutions is significantly greater than the rate of non-synonymous substitutions between E. tallapoosae and the two Percina species (p < 0.05). The remainder of the sequence is under purifying selection (data not shown).

Bottom Line: As a result of this study we have demonstrated that the Tallapoosa darter CENP-A gene HFD sequences can be isolated from genomic DNA by nested PCR in a manner that does not lead to the amplification of the highly sequence related histone H3 gene.These adjacent gene sequences provide us with a primer binding sites for PCR isolation of the CENP-A gene from other percid species of fishes.An initial comparison of three percid species shows that the N-terminal tail of the percid CENP-A gene shows adaptive evolution.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of West Georgia, Carrollton, GA 30118, USA. dfountain82@gmail.com.

ABSTRACT
Both centromeric alpha-satellite sequences as well as centromeric protein A (CENP-A) are highly variable in eukaryotes. CENP-A, a histone H3 variant, is thought to act as the epigenetic "mark" for assembly of centromeric proteins. While most of the histone fold domain (HFD) of the CENP-A is fairly well conserved, a portion of this HFD as well as the N-terminal tail show adaptive variation in both plants and animals. Such variation may establish reproductive barriers that may lead to speciation. The family Percidae contains over 200 species most of which are within the subfamily Etheostomatinae. This subfamily represents a species rich radiation of freshwater fishes in North America and these species exhibit both allopatric and sympatric distributions. In order to study the evolution of CENP-A in percid fish species, we have isolated and characterized the CENP-A gene from Etheostoma tallapoosae by PCR based gene walking. As a result of this study we have demonstrated that the Tallapoosa darter CENP-A gene HFD sequences can be isolated from genomic DNA by nested PCR in a manner that does not lead to the amplification of the highly sequence related histone H3 gene. We also demonstrated that PCR based walking can be subsequently used to isolate the rest of the CENP-A gene and adjacent gene sequences. These adjacent gene sequences provide us with a primer binding sites for PCR isolation of the CENP-A gene from other percid species of fishes. An initial comparison of three percid species shows that the N-terminal tail of the percid CENP-A gene shows adaptive evolution.

No MeSH data available.


Related in: MedlinePlus