Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya
Papaya is a globally important crop, with production primarily based on hermaphrodite plants. Papaya has three sex types—male, female, and hermaphrodite—determined by flower morphology, but this is only distinguishable at the flowering stage. In this study, a recombinase polymerase amplification (RP...
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MDPI AG
2024-09-01
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| Online Access: | https://www.mdpi.com/2813-0464/3/4/27 |
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| author | José Guadalupe Ávila-Hernández Alejandro Coreño-Alonso Mario Alberto Pantoja-Alonso Francisco Javier Córdoba-Andrade Rogelio González-González Corina E. Díaz-Quezada Alberto Camas-Reyes Agustino Martínez-Antonio |
| author_facet | José Guadalupe Ávila-Hernández Alejandro Coreño-Alonso Mario Alberto Pantoja-Alonso Francisco Javier Córdoba-Andrade Rogelio González-González Corina E. Díaz-Quezada Alberto Camas-Reyes Agustino Martínez-Antonio |
| author_sort | José Guadalupe Ávila-Hernández |
| collection | DOAJ |
| description | Papaya is a globally important crop, with production primarily based on hermaphrodite plants. Papaya has three sex types—male, female, and hermaphrodite—determined by flower morphology, but this is only distinguishable at the flowering stage. In this study, a recombinase polymerase amplification (RPA) assay was developed and optimized to identify the three sexes of papaya. Recombinant uvsX, uvsY, gp32, and Bsu DNA polymerase were used to study the effects of temperature, reaction time, and sensitivity conditions for RPA reaction efficiency. The optimal conditions were found to be 41 °C and a 30 min reaction time, allowing the detection of the target sex from specific DNA markers, even when using crude extract. This study shows that RPA could be used for sex determination in papaya, and the findings could contribute to developing a point-of-need strategy due to their sensitivity and specificity. |
| format | Article |
| id | doaj-art-c36019d2492943cc8a5db402a467fa97 |
| institution | DOAJ |
| issn | 2813-0464 |
| language | English |
| publishDate | 2024-09-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Applied Biosciences |
| spelling | doaj-art-c36019d2492943cc8a5db402a467fa972025-08-20T02:53:27ZengMDPI AGApplied Biosciences2813-04642024-09-013442643710.3390/applbiosci3040027Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in PapayaJosé Guadalupe Ávila-Hernández0Alejandro Coreño-Alonso1Mario Alberto Pantoja-Alonso2Francisco Javier Córdoba-Andrade3Rogelio González-González4Corina E. Díaz-Quezada5Alberto Camas-Reyes6Agustino Martínez-Antonio7Departamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoDepartamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoDepartamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoDepartamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoUnidad de Genómica Avanzada, Centro de Investigación y de Estudios Avanzados, Irapuato 36824, Guanajuato, MexicoUnidad de Genómica Avanzada, Centro de Investigación y de Estudios Avanzados, Irapuato 36824, Guanajuato, MexicoDepartamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoDepartamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados (CINVESTAV-IPN), Unidad Irapuato, Irapuato 36824, Guanajuato, MexicoPapaya is a globally important crop, with production primarily based on hermaphrodite plants. Papaya has three sex types—male, female, and hermaphrodite—determined by flower morphology, but this is only distinguishable at the flowering stage. In this study, a recombinase polymerase amplification (RPA) assay was developed and optimized to identify the three sexes of papaya. Recombinant uvsX, uvsY, gp32, and Bsu DNA polymerase were used to study the effects of temperature, reaction time, and sensitivity conditions for RPA reaction efficiency. The optimal conditions were found to be 41 °C and a 30 min reaction time, allowing the detection of the target sex from specific DNA markers, even when using crude extract. This study shows that RPA could be used for sex determination in papaya, and the findings could contribute to developing a point-of-need strategy due to their sensitivity and specificity.https://www.mdpi.com/2813-0464/3/4/27recombinase polymerase amplification<i>Carica papaya</i> L.isothermal amplificationmolecular sexinghermaphrodite |
| spellingShingle | José Guadalupe Ávila-Hernández Alejandro Coreño-Alonso Mario Alberto Pantoja-Alonso Francisco Javier Córdoba-Andrade Rogelio González-González Corina E. Díaz-Quezada Alberto Camas-Reyes Agustino Martínez-Antonio Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya Applied Biosciences recombinase polymerase amplification <i>Carica papaya</i> L. isothermal amplification molecular sexing hermaphrodite |
| title | Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya |
| title_full | Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya |
| title_fullStr | Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya |
| title_full_unstemmed | Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya |
| title_short | Step-by-Step Development of a Recombinase Polymerase Amplification (RPA) Assay for Sex Identification in Papaya |
| title_sort | step by step development of a recombinase polymerase amplification rpa assay for sex identification in papaya |
| topic | recombinase polymerase amplification <i>Carica papaya</i> L. isothermal amplification molecular sexing hermaphrodite |
| url | https://www.mdpi.com/2813-0464/3/4/27 |
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