RESUMO
FLORICAULA/LEAFY (FLO/LFY) is a single-copy nuclear-encoded homeotic gene containing two introns. We have investigated the utility of the second intron of FLO/LFY (FLint2) as a tool for phylogeny reconstruction at lower taxonomic levels. As an example, the phylogeny of 46 Amorphophallus, two Pseudodracontium, and four outgroup species is reconstructed using maximum parsimony and maximum likelihood analyses of FLint2 sequences. We designed new primers based on conserved sequences of the second and third exon for use in a range of Aroid taxa to amplify and sequence the second intron. In Amorphophallus FLint2 proved to be rather short (143-222 bp), highly variable and unsaturated. In all but two species a single amplification product was found. Results from phylogenetic analysis of FLint2 are largely congruent with results using the chloroplast regions rbcL, matK, and trnL, and compare favorably in percentage of informative characters, overall homoplasy levels, number of well-supported clades in consensus trees and resolution of ingroup relationships within Amorphophallus. When amplification products are not too large, alignment is relatively straightforward, and sequences are used in combination with other fast evolving markers, the FLint2 intron may be a valuable new tool for phylogenetic studies at lower taxonomic levels.
Assuntos
Amorphophallus/classificação , Amorphophallus/genética , Cloroplastos/genética , DNA de Plantas/genética , Filogenia , Sequência de Bases , Primers do DNA , Íntrons , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
Late-infantile neuronal ceroid lipofuscinosis (LINCL) is a progressive neurodegenerative disorder caused by the deficiency of lysosomal tripeptidyl peptidase I (TPP-I) encoded by the CLN2 gene. We report the first case of early prenatal diagnosis of LINCL by combined enzyme and mutation analysis. TPP-I activity in chorionic villi (CV) was less than 2% of the mean normal control level and g.1946A > G and g.3670C > T mutations were demonstrated, as in the two previously affected children. After termination of pregnancy, TPP-I deficiency was confirmed in cultured CV cells and in the fetal skin fibroblasts. The expression of unequivocal TPP-I deficiency in CV demonstrates that enzyme assay is a reliable option for prenatal diagnosis of LINCL.
Assuntos
Análise Mutacional de DNA , Endopeptidases/deficiência , Endopeptidases/genética , Lipofuscinoses Ceroides Neuronais/diagnóstico , Diagnóstico Pré-Natal , Aminopeptidases , Vilosidades Coriônicas/enzimologia , Amostra da Vilosidade Coriônica , Dipeptidil Peptidases e Tripeptidil Peptidases , Feminino , Humanos , Lipofuscinoses Ceroides Neuronais/enzimologia , Lipofuscinoses Ceroides Neuronais/genética , Gravidez , Primeiro Trimestre da Gravidez , Tripeptidil-Peptidase 1Assuntos
Deleção Cromossômica , Cromossomo Y , Guias como Assunto , Humanos , Laboratórios , Controle de QualidadeRESUMO
Infantile neuronal ceroid lipofuscinosis (INCL) is a progressive neurodegenerative disorder in childhood which is caused by the deficiency of the lysosomal palmitoyl-protein thioesterase (PPT) encoded by the CLN1 gene. In a pregnancy at risk for INCL, chorionic villi (CV) were studied using a novel fluorometric PPT enzyme assay in combination with mutation-analysis of the CLN1 gene. The PPT activity in chorionic villi was found to be deficient and homozygosity for the C451T mutation in CLN1 was found. The pregnancy was terminated and the PPT deficiency was confirmed in cultured CV cells as well as in the cultured fetal skin fibroblasts. This report shows the first early prenatal diagnosis of INCL performed by fluorometric enzyme analysis and mutation analysis of the CLN1 gene.
