An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding
In applications such as marker-assisted breeding and positional cloning, tissue sampling and plant tracking are vital steps in the genotyping pipeline. They enable the identification of desirable seedlings, saving time and reducing the cost, space, and handling required for growing adult plants, esp...
Saved in:
| Main Authors: | , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Bio-protocol LLC
2025-02-01
|
| Series: | Bio-Protocol |
| Online Access: | https://bio-protocol.org/en/bpdetail?id=5200&type=0 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1825206638878916608 |
|---|---|
| author | Brian Zebosi John Ssengo Lander Geadelmann Erica Unger-Wallace Erik Vollbrecht |
| author_facet | Brian Zebosi John Ssengo Lander Geadelmann Erica Unger-Wallace Erik Vollbrecht |
| author_sort | Brian Zebosi |
| collection | DOAJ |
| description | In applications such as marker-assisted breeding and positional cloning, tissue sampling and plant tracking are vital steps in the genotyping pipeline. They enable the identification of desirable seedlings, saving time and reducing the cost, space, and handling required for growing adult plants, especially for greenhouses and winter nurseries. Small-scale marker-assisted selection laboratories rely heavily on leaf-based genotyping, which involves over-planting large, segregating populations followed by leaf sampling, genotyping, and backtracking to identify desired individuals, which is costly and laborious. Thus, there is a need to adopt seed-based genotyping to reduce costs and save time. Therefore, we developed a safe and cheap seed-chipping protocol using clipping pliers to chip seeds to genotype before planting. To identify a cost-effective and high-throughput DNA extraction method, we tested four extraction methods and assessed the quality of the seed DNA using PCR. For three of the methods, seed-based DNA was of comparable quality to DNA extracted from leaf punches. We also compared seed- and leaf-derived DNA from the same individuals in a segregating population to test for genotyping miscalls that could arise due to the presence of maternally derived pericarp in the seed samples. Out of 43 potential instances, we found zero miscalled samples and, therefore, no evidence supporting consequential pericarp inclusion. Germination rates of chipped and unchipped seeds were the same for the inbreds tested, B73 and Mo17. However, chipped seeds grew slower until ~14 days after sowing. Overall, seed sampling using clipping pliers provides a simple, reliable, and high-throughput method to identify specific genotypes before planting. |
| format | Article |
| id | doaj-art-a6c78d643d7c40f28181c6787cae88e9 |
| institution | Kabale University |
| issn | 2331-8325 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Bio-protocol LLC |
| record_format | Article |
| series | Bio-Protocol |
| spelling | doaj-art-a6c78d643d7c40f28181c6787cae88e92025-02-07T08:16:46ZengBio-protocol LLCBio-Protocol2331-83252025-02-0115310.21769/BioProtoc.5200An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted BreedingBrian Zebosi0John Ssengo1Lander Geadelmann2Erica Unger-Wallace3Erik Vollbrecht4Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, USAGenetics and Genomics Graduate Program, Iowa State University, Ames, IA, USAGenetics and Genomics Graduate Program, Iowa State University, Ames, IA, USADepartment of Plant Pathology, Entomology and Microbiology, Iowa State University, Ames, IA, USADepartment of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, USAPlant Biology Graduate Program, Iowa State University, Ames, IA, USADepartment of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, USADepartment of Genetics, Development and Cell Biology, Iowa State University, Ames, IA, USAGenetics and Genomics Graduate Program, Iowa State University, Ames, IA, USA, Plant Biology Graduate Program, Iowa State University, Ames, IA, USAIn applications such as marker-assisted breeding and positional cloning, tissue sampling and plant tracking are vital steps in the genotyping pipeline. They enable the identification of desirable seedlings, saving time and reducing the cost, space, and handling required for growing adult plants, especially for greenhouses and winter nurseries. Small-scale marker-assisted selection laboratories rely heavily on leaf-based genotyping, which involves over-planting large, segregating populations followed by leaf sampling, genotyping, and backtracking to identify desired individuals, which is costly and laborious. Thus, there is a need to adopt seed-based genotyping to reduce costs and save time. Therefore, we developed a safe and cheap seed-chipping protocol using clipping pliers to chip seeds to genotype before planting. To identify a cost-effective and high-throughput DNA extraction method, we tested four extraction methods and assessed the quality of the seed DNA using PCR. For three of the methods, seed-based DNA was of comparable quality to DNA extracted from leaf punches. We also compared seed- and leaf-derived DNA from the same individuals in a segregating population to test for genotyping miscalls that could arise due to the presence of maternally derived pericarp in the seed samples. Out of 43 potential instances, we found zero miscalled samples and, therefore, no evidence supporting consequential pericarp inclusion. Germination rates of chipped and unchipped seeds were the same for the inbreds tested, B73 and Mo17. However, chipped seeds grew slower until ~14 days after sowing. Overall, seed sampling using clipping pliers provides a simple, reliable, and high-throughput method to identify specific genotypes before planting.https://bio-protocol.org/en/bpdetail?id=5200&type=0 |
| spellingShingle | Brian Zebosi John Ssengo Lander Geadelmann Erica Unger-Wallace Erik Vollbrecht An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding Bio-Protocol |
| title | An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding |
| title_full | An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding |
| title_fullStr | An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding |
| title_full_unstemmed | An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding |
| title_short | An Effective and Safe Maize Seed Chipping Protocol Using Clipping Pliers With Applications in Small-Scale Genotyping and Marker-Assisted Breeding |
| title_sort | effective and safe maize seed chipping protocol using clipping pliers with applications in small scale genotyping and marker assisted breeding |
| url | https://bio-protocol.org/en/bpdetail?id=5200&type=0 |
| work_keys_str_mv | AT brianzebosi aneffectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT johnssengo aneffectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT landergeadelmann aneffectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT ericaungerwallace aneffectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT erikvollbrecht aneffectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT brianzebosi effectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT johnssengo effectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT landergeadelmann effectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT ericaungerwallace effectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding AT erikvollbrecht effectiveandsafemaizeseedchippingprotocolusingclippingplierswithapplicationsinsmallscalegenotypingandmarkerassistedbreeding |