Source: AGRICULTURAL RESEARCH SERVICE submitted to
GENETIC ANALYSES OF A CHLOROPLAST-ASSOCIATED COLD TOLERANCE
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
NEW
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1008493
Grant No.
2016-67013-24590
Project No.
ILLW-2015-06563
Proposal No.
2015-06563
Multistate No.
(N/A)
Program Code
A1101
Project Start Date
Dec 15, 2015
Project End Date
Dec 14, 2019
Grant Year
2016
Project Director
Havey, M. J.
Recipient Organization
AGRICULTURAL RESEARCH SERVICE
1815 N University
Peoria,IL 61604
Performing Department
Department of Horticulture
Non Technical Summary
Warm-season crops are severely damaged by cold stress. Growers would benefit from cold-tolerant cultivars which can be planted earlier in the spring, survive short periods of cold temperatures, and be harvested before diseases and pests reach their greatest threat later in the season. Cucumber is extremely susceptible to cold stress and a maternally inherited cold tolerance has been identified. This research will exploit the unique transmission modes of the cucumber organelles (maternal chloroplast and paternal mitochondrion) to identify differences in the chloroplast DNA, RNA, and proteins differing between cold-susceptible versus tolerant chloroplasts. We will cross among doubled haploids of diverse cucumbers to produce cold-susceptible or tolerant reciprocal hybrids with identical nuclear genotypes, and grow reciprocal hybrids and parents in replicated trials in optimal and cold-stress environments. Sequencing of the chloroplast DNAs and RNAs will be completed to identify differences that distinguish cold-susceptible from tolerant chloroplasts. Targeted analyses of the chloroplast proteins will be used to confirm predicted differences from the DNA and/or RNA. The chloroplast-associated cold tolerance will be backcrossed into elite cucumber inbreds for release to stakeholders. These studies should provide deeper insights about organellar differences that can be selected to develop cold-tolerant crops to enhance the sustainability of US production agriculture under challenging and changing climatic conditions.
Animal Health Component
0%
Research Effort Categories
Basic
50%
Applied
50%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2011421108125%
2031421108075%
Knowledge Area
203 - Plant Biological Efficiency and Abiotic Stresses Affecting Plants; 201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1421 - Cucumber;

Field Of Science
1080 - Genetics; 1081 - Breeding;
Goals / Objectives
Warm-season crops are severely damaged by cold stress. Growers would benefit from cold-tolerant cultivars which can be planted earlier in the spring, survive short periods of cold temperatures, and be harvested before diseases and pests reach their greatest threat later in the season. Cucumber is extremely susceptible to cold stress and a maternally inherited cold tolerance has been identified. This cold tolerance is associated with maternally transmitted chloroplasts because the mitochondria of cucumber show paternal transmission. We will exploit these unique transmission modes to identify differences in the genome, transcriptome, and proteome of cold-susceptible versus tolerant chloroplasts.
Project Methods
We crossed among doubled haploids of diverse cucumbers to produce cold-susceptible or tolerant reciprocal hybrids with identical nuclear genotypes. We will grow reciprocal hybrids and parents in replicated trials in optimal and cold-stress environments. Next-gen sequencing of the chloroplast DNAs and RNAs will be completed to determine if heteroplasmy, gene-expression differences, or post-transcriptional modifications distinguish cold-susceptible from tolerant chloroplasts. Targeted analyses of the chloroplast proteome will be used to confirm predicted differences from the genome and/or transcriptome. Backcrossing of the cold-tolerant chloroplasts into elite cucumber inbreds will be completed in the greenhouse by standard crossing techniques.

Progress 12/15/16 to 12/14/17

Outputs
Target Audience: Nothing Reported Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Graduate student research training. The student will present a poster at the Plant and Animal Genome Conference in 2018 How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?In the next year, we will focus on RNA analyses of chloroplast transcriptsof cold-tolerant and susceptible cucumber lines. We will determine the level and postions of RNA editing to establish if any differences occur after editing. We will also determine if the candidate SNP in the chloroplast DNA is edited.

Impacts
What was accomplished under these goals? Chloroplast DNAs were purified from cold-tolerant and susceptible lines of cucumber. The cold-tolerant line has a single nucleotide polymorphism (SNP) in one of the subunits of atpase, which results in an amino acid change. This specific SNP is the candidate for cold tolerance.

Publications


    Progress 12/15/15 to 12/14/16

    Outputs
    Target Audience:Cucumber breeders and producers. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?A graduate student was hired and she started on September 1, 2016. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?We plan to complete next-generation sequencing of chloroplast DNAs from cold-tolerant and susceptible doubled-haploid lines, and evaluate for sublimons that may be associated with cold tolerance.

    Impacts
    What was accomplished under these goals? Doubled haploid (DH) cucumber lines were produced and evaluated under cold stress. The DH from the heirloom cultivar 'Chipper' suffered significantly less damage by cold compared to other DH lines from cultivars 'Straight 8' and '9930'. Hybrids were made between the Chipper DH with the cold-susceptible DHs and evaluated for cold tolerance. The hybrids with Chipper as the maternal parent performed significantly better than the reciprocal hybrids. Chloroplast DNA was isolated from DHs of Chipper and Straight 8 for next-generation sequencing.

    Publications