Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA)
Single strand conformation analysis (SSCA) is a technique that has been used to detect point mutations. We explored its usefulness in the analysis of four different members of the Trypanosoma cruzi TcP2β gene family and its suitability for detection of polymorphism in different parasite strains. The...
Guardado en:
| Publicado: |
1996
|
|---|---|
| Materias: | |
| Acceso en línea: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03781119_v180_n1-2_p43_Vazquez http://hdl.handle.net/20.500.12110/paper_03781119_v180_n1-2_p43_Vazquez |
| Aporte de: |
| id |
paper:paper_03781119_v180_n1-2_p43_Vazquez |
|---|---|
| record_format |
dspace |
| spelling |
paper:paper_03781119_v180_n1-2_p43_Vazquez2023-06-08T15:39:14Z Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) Protozoan parasites genome projects Ribosomal P protein genes Sequence Tagged Sites Trypanosoma cruzi glycerol glycine cleavage system polyacrylamide gel ribosome protein article controlled study dna polymorphism electrophoretic mobility gene amplification genome marker gene multigene family nonhuman nucleic acid base substitution point mutation polymerase chain reaction priority journal single strand conformation polymorphism trypanosoma cruzi Animals Base Sequence Genes, Protozoan Molecular Sequence Data Multigene Family Phosphoproteins Polymerase Chain Reaction Polymorphism, Single-Stranded Conformational Protozoan Proteins Ribosomal Proteins Sequence Tagged Sites Trypanosoma cruzi Protozoa Trypanosoma Trypanosoma cruzi Single strand conformation analysis (SSCA) is a technique that has been used to detect point mutations. We explored its usefulness in the analysis of four different members of the Trypanosoma cruzi TcP2β gene family and its suitability for detection of polymorphism in different parasite strains. The availability of primers covering a 97-bp sequence at the 5' end of the genes allowed assessment of the effect of a single base substitution, while the analysis of a 321 bp long sequence permitted the evaluation of sequences differing in several bases. PCR products were analysed under four different electrophoretic conditions: with or without the addition of 10% glycerol in a 6% polyacrylamide gel run at room temperature or at 4°C. Shifts in mobility were radically dependent on the migration condition. Both 97-bp and 321-bp amplicons were best resolved at 4°C, without glycerol. Amplification products derived from total genomic DNA showed a pattern that resembled closely a combination of the products derived from the cloned genes. The results herein demonstrate the usefulness of SSCA to differentiate forms of a complex protozoan gene family, and to scan its polymorphic nature. Furthermore, due to the remarkable sensitivity of the technique it can generate genomic markers, such as Sequence Tagged Sites (STS), of great need in the T. cruzi genome project. 1996 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03781119_v180_n1-2_p43_Vazquez http://hdl.handle.net/20.500.12110/paper_03781119_v180_n1-2_p43_Vazquez |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Protozoan parasites genome projects Ribosomal P protein genes Sequence Tagged Sites Trypanosoma cruzi glycerol glycine cleavage system polyacrylamide gel ribosome protein article controlled study dna polymorphism electrophoretic mobility gene amplification genome marker gene multigene family nonhuman nucleic acid base substitution point mutation polymerase chain reaction priority journal single strand conformation polymorphism trypanosoma cruzi Animals Base Sequence Genes, Protozoan Molecular Sequence Data Multigene Family Phosphoproteins Polymerase Chain Reaction Polymorphism, Single-Stranded Conformational Protozoan Proteins Ribosomal Proteins Sequence Tagged Sites Trypanosoma cruzi Protozoa Trypanosoma Trypanosoma cruzi |
| spellingShingle |
Protozoan parasites genome projects Ribosomal P protein genes Sequence Tagged Sites Trypanosoma cruzi glycerol glycine cleavage system polyacrylamide gel ribosome protein article controlled study dna polymorphism electrophoretic mobility gene amplification genome marker gene multigene family nonhuman nucleic acid base substitution point mutation polymerase chain reaction priority journal single strand conformation polymorphism trypanosoma cruzi Animals Base Sequence Genes, Protozoan Molecular Sequence Data Multigene Family Phosphoproteins Polymerase Chain Reaction Polymorphism, Single-Stranded Conformational Protozoan Proteins Ribosomal Proteins Sequence Tagged Sites Trypanosoma cruzi Protozoa Trypanosoma Trypanosoma cruzi Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| topic_facet |
Protozoan parasites genome projects Ribosomal P protein genes Sequence Tagged Sites Trypanosoma cruzi glycerol glycine cleavage system polyacrylamide gel ribosome protein article controlled study dna polymorphism electrophoretic mobility gene amplification genome marker gene multigene family nonhuman nucleic acid base substitution point mutation polymerase chain reaction priority journal single strand conformation polymorphism trypanosoma cruzi Animals Base Sequence Genes, Protozoan Molecular Sequence Data Multigene Family Phosphoproteins Polymerase Chain Reaction Polymorphism, Single-Stranded Conformational Protozoan Proteins Ribosomal Proteins Sequence Tagged Sites Trypanosoma cruzi Protozoa Trypanosoma Trypanosoma cruzi |
| description |
Single strand conformation analysis (SSCA) is a technique that has been used to detect point mutations. We explored its usefulness in the analysis of four different members of the Trypanosoma cruzi TcP2β gene family and its suitability for detection of polymorphism in different parasite strains. The availability of primers covering a 97-bp sequence at the 5' end of the genes allowed assessment of the effect of a single base substitution, while the analysis of a 321 bp long sequence permitted the evaluation of sequences differing in several bases. PCR products were analysed under four different electrophoretic conditions: with or without the addition of 10% glycerol in a 6% polyacrylamide gel run at room temperature or at 4°C. Shifts in mobility were radically dependent on the migration condition. Both 97-bp and 321-bp amplicons were best resolved at 4°C, without glycerol. Amplification products derived from total genomic DNA showed a pattern that resembled closely a combination of the products derived from the cloned genes. The results herein demonstrate the usefulness of SSCA to differentiate forms of a complex protozoan gene family, and to scan its polymorphic nature. Furthermore, due to the remarkable sensitivity of the technique it can generate genomic markers, such as Sequence Tagged Sites (STS), of great need in the T. cruzi genome project. |
| title |
Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| title_short |
Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| title_full |
Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| title_fullStr |
Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| title_full_unstemmed |
Detection of polymorphism in the Trypanosoma cruzi TcP2β gene family by single strand conformational analysis (SSCA) |
| title_sort |
detection of polymorphism in the trypanosoma cruzi tcp2β gene family by single strand conformational analysis (ssca) |
| publishDate |
1996 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_03781119_v180_n1-2_p43_Vazquez http://hdl.handle.net/20.500.12110/paper_03781119_v180_n1-2_p43_Vazquez |
| _version_ |
1768542367033327616 |