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Molecular basis underlying the QTL responsible for the genetic control of flowering time in chickpea: an integrative approach

Periodic Reporting for period 1 - FORCE (Molecular basis underlying the QTL responsible for the genetic control of flowering time in chickpea: an integrative approach)

Reporting period: 2019-10-01 to 2021-09-30

Chickpea is an annual, self-pollinated diploid crop species with a genome size of ~ 740 Mb. Globally, chickpea is the third most important food legume. Because of the high protein content of the seeds, chickpea is considered a stable protein source for the human diet. As a legume, chickpea contributes to the sustainable improvement of the environment when grown in agricultural rotations due to its ability to fix nitrogen.

Chickpea underwent domestication (the transition from wild to crop species) as part of the evolution of agriculture in the Fertile Crescent 12,000 to 10,000 years ago. Following domestication, chickpea cultivation expanded to wider latitudinal and climatic range with shorter growing seasons where the plant life cycle can be completed before the end of summer, permitting the escape from end-of-season stresses that constrain stable production​​. However, domestication and artificial selection resulted in a very ​narrow genetic diversity of modern cultivated chickpea germplasm, and breeding programs are challenged by this evolutionary bottleneck.

Flowering time involves the transition from vegetative growth to the flowering state and it refers to the number of days from sowing to the appearance of the first fully opened flower​. Flowering time is the ​major domestication trait defining the adaptation of chickpea to different agro-climatic conditions, and therefore is a major determinant of its ​productivity​.
Despite early flowering time being suggested as a means to increase chickpea adaptability roughly 45 years ago ​, studies on the genetic control of flowering time have appeared only within the 21st century. Although classical genetic analyses have provided a wealth of valuable information, the molecular identities of genes underlying the major flowering time locus remains mostly elusive.
Understanding and learning the allelic variation of the complex gene network controlling flowering time will allow us to obtain molecular tags for early/late flowering tags that may ultimately be used in the breeding programs to develop adapted cultivars to their specific environments. This could be exploited by programs of international institutions (ICARDA, ICRISAT, INRAT) and private companies. This Marie Skłodowska Curie Action (MSCA) emphasizes the use of ​modern methodologies in genomics and bioinformatics, together with plant breeding​. This MSCA links to the Horizon 2020 Programme of the EU under the specific challenge ​Food security, sustainable agriculture and forestry research (Work Programme 2018-2020), specifically through the innovations in plant variety testing​ section: “identify crop characteristics with the capacity ... to maintain yield under more variable conditions and under more sustainable crop management practices”.
Formal objectives of this Marie Skłodowska Curie Action (MSCA) have been to (a) identify ​the molecular basis underlying the genomic regions responsible for the genetic control of flowering time ​in chickpea; (b) develop molecular markers for early/late flowering
The WP3 has been the most time-consuming activity of the Project. WP3 involves seed preparation, growing chambers setup, monitorization of emergency days, and phenotypic evaluation for determining the day of flowering for each plant.
So far, we have collected data for 5 recombinant inbred lines involving > 2,500 plants with an averaged stay of 200 days in the growth chamber per population. The contrasting genotypes in each population plus their parents have been sequenced (~400 plants) and the following workflow has been implemented for each population raw data :
* quality trimming to remove low quality regions
* alignment to the reference genome
* SNP calling (homozygous SNPs and heterozygous SNPs)

At the time of this report, we have identified a set of ~5,000 - 8,000 high-quality SNPs per population from > 725,000,000 raw reads.
With > 20,800 high-quality SNPs, population RIP12 is particularly interesting as it involves an interspecific cross using the wild species Cicer reticulatum as a parent. C. reticulatum is considered to be the ancestor of the cultivated chickpea. As expected, the population structure of the lines from that interspecific cross is quite different compared to the other populations.
The genotyping and sequence analyses are currently underway and another population is under evaluation at the time of this writing. The WP3 represents the core activity of the MSCA project and its completion will allow us to construct the highest-density genetic map available for flowering time in chickpea using large-scale mapping populations. Our results will be published in a scientific journal paper.

The communication strategy in our project involved the dissemination of results. So far, we have achieved this (and we continue working on it) through attendance at meetings, use of social media (twiter), the creation of a website, and collaborations with Marie Curie Alumni Association, among others.
As stated above, we are currently generating a vast amount of data based on genome sequences from plant individuals that have been already phenotyped. This represents the core of the project and is ongoing work. As the University of Cordoba has chickpea populations coming from different genetic backgrounds , we expect to find the early flowering trait being controlled not necessarily by the same molecular mechanism. In each population, the effect of temperature, photoperiod, and vernalization may be different and our integrative approach should be able to understand those environmental triggers and the further influence on the flowering adaptive trait.
Our ultimate goal by understanding the allelic variation of the gene network controlling flowering time is to develop molecular markers for flowering that will ultimately be used by the breeding programs to develop adapted cultivars to their specific environments.