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Mendez, M.; Parera, V.E.; Rossetti, M.V.; De Siervi, A.; Astrin, K.H.; Desnick, J.R.; Del C Batlle, A.M. "Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa" (1998) Revista Argentina de Dermatologia. 79(1):20-30
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Abstract:

Porphyria Cutanea Tarda (PCT) and Hepatoerythropoietic Porphyria (HEP) are characterized by a deficiency in the activity of Uroporphyrinogen decarboxyiase (URO-D), the fifth enzyme in the heme pathway, which catalises the sequential decarboxylation of Uroporphyrinogen to Coproporphyrinogen.There are two types of PCT: sporadic (S-PCT) and familial (F-PCT).The former is an acquired disorder, whereby URO-D activity is only diminished in the liver while the latter is a hereditary dominant disorder where URO-D activity is 50% reduced in all tissues. HEP is a recessive disease where URO-D activity is diminished to 5-10% of normal value. In both porphyrias accumulation of highly carboxilated porphyrins produces the characteristic cutaneous photosensitivity. Human URO-D gene maps in chromosomel (1p34) and comprises 10 exons spread on 3 kb. UROD mRNA has 1,197 bases which codifies a protein of 367 aminoacids.Ten mutations in URO-D gene have been already identified causing F-PCT or HEP. In this work we describe the case of an argentinean patient with the classical symptoms of PCT: high urinary porphyrins levels (8,909; NV=20-250 μg/24 hrs), with the characteristic excretion pattern and an elevated plasma porphyrin index (7.48; NV < 1.30). Erythrocytic URO-D activity was measured employing Uroporphyrinogen as substrate and the porphyrins obtained were analized by HPLC. The URO-D value of the patient was 7.33 U/ml RBC (45% of normal value= 16.35 ±3.56 U/ml RBC). Lymphocytes were separated first from fesh whole blood by density gradient, total RNA was isolated using the commercial reagent RNA zol ™B (BIOTECX). Then RT-PCR reactions were carried out employing M-MLV RT and oligo(dT) for RT andTaq DNA polymerase and the adequate primers for amplification. The product was purified employing Wizard™ (PROMEGA) minicolums and it was sequenced by Sanger methodology using a modified Taq polymerase (fmol PROMEGA) or sequence after asymétrie PCR to obtain both single strands separately. Two products were obtained in the RT-PCR; both bands were cut from 2% agarose gel and eluted in TE buffer. They were then sequenced, showing that in the small band the 67 bases corresponding to exon 9 were missing. DNA genomic analysis showed the transition G→A in the last base of exon 9 (E314E) causing its skipping. Furthermore, due to the joint of exons 8 and 10 the reading frame was changed, leading to the formation of a stop codon and the synthesis of a truncate protein with the losing of about 20% of its aminoacids.

Registro:

Documento: Artículo
Título:Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa
Autor:Mendez, M.; Parera, V.E.; Rossetti, M.V.; De Siervi, A.; Astrin, K.H.; Desnick, J.R.; Del C Batlle, A.M.
Filiación:Centro de Investigaciones Sobre Porfirinas y Porfirias (CIPYP), Facultad de Ciencias Exactas y Naturales, UBA - Consejo Nacional de Investigaciones Cientificas y Técnicas (CONICET), Buenos Aires, Argentina
Departamento de Genética Humana, Mount Sinai School of Medicine, Nueva York, EE.UU, United States
Becario de Perfeccionainienlo (CONICET), Argentina
Departamento de Genética Humana, Mount Sinai School of Medicine, Nueva York, United States
Año:1998
Volumen:79
Número:1
Página de inicio:20
Página de fin:30
Título revista:Revista Argentina de Dermatologia
Título revista abreviado:Rev. Argent. Dermatol.
ISSN:03252787
CODEN:RADEB
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03252787_v79_n1_p20_Mendez

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Citas:

---------- APA ----------
Mendez, M., Parera, V.E., Rossetti, M.V., De Siervi, A., Astrin, K.H., Desnick, J.R. & Del C Batlle, A.M. (1998) . Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa. Revista Argentina de Dermatologia, 79(1), 20-30.
Recuperado de https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03252787_v79_n1_p20_Mendez [ ]
---------- CHICAGO ----------
Mendez, M., Parera, V.E., Rossetti, M.V., De Siervi, A., Astrin, K.H., Desnick, J.R., et al. "Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa" . Revista Argentina de Dermatologia 79, no. 1 (1998) : 20-30.
Recuperado de https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03252787_v79_n1_p20_Mendez [ ]
---------- MLA ----------
Mendez, M., Parera, V.E., Rossetti, M.V., De Siervi, A., Astrin, K.H., Desnick, J.R., et al. "Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa" . Revista Argentina de Dermatologia, vol. 79, no. 1, 1998, pp. 20-30.
Recuperado de https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03252787_v79_n1_p20_Mendez [ ]
---------- VANCOUVER ----------
Mendez, M., Parera, V.E., Rossetti, M.V., De Siervi, A., Astrin, K.H., Desnick, J.R., et al. Porfiria cutanea tardia: Deteccion de una nueva mutacion en la uroporfirinogeno decarboxilasa mediante rt-pcr y secuenciacion directa. Rev. Argent. Dermatol. 1998;79(1):20-30.
Available from: https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03252787_v79_n1_p20_Mendez [ ]