Genetic Architecture Of Nodal Root Number During Domestication And Cloning Of Stiff Stalk Gene Stiff1 In Maize(Zea Mays L.)

Maize(Zea mays L.)was domesticated from its progenitor teosinte(Zea mays.parviglumis)approximately 10,000 years ago.As a typical high yielding C4 cereal,maize plays an important role in global food security providing more than 30%of food caloric for human.Maize has been planting in the largest area in the world compared with other cereals since last decade.The maize nodal root system plays a crucial role in the development of the aboveground plant and determines the yield via the uptake of water and nutrients in the field.Meanwhile,stalk lodging is one main threat to high yield.And stiff stalk is also an important trait pursed in miaze modern dense-planting breeding.Not only will strong stalk enhance the facility of mechanical grain harvesting,but also the tolerance of biotic and abiotic stresses originated from high-density planting in maize.However,the genetic architectures of maize root system and stiff stalk are not well understood because of the complex genetic background and the difficulties of measurement.In this study,the genetic mechanisms of maize nodal root and a major quantitative trait locus(QTL)controlling stiff stalk were analyzed.The major results are described as follows:1.Through a large teosinte-maize population,high-resolution mapping revealed that 62 out of 133 QTLs.accounting for approximately half of the total genetic variation in nodal root number,were derived from QTLs for flowering time.Whereas only 16%of the total genetic variation in nodal root number was derived from QTLs for plant height.2.The QTL peaks of the crown nodal root number traits were colocalized with the major QTL-ZmCCT for flowering time on chromosome 10.Comparison of the underground nodal root number traits including CRN,CRLN and CRNPL between ZmCCT near-isogenic lines(NILs)and between ZmCCT transgenic and non-transgenic control plants suggested that the ZmCCT co-controlled the nodal root number and flowering time.In addition,3 more candidate genes governed both the nodal root number and flowering time were identified:GRMZM2G133555(CCT)on chromosome 1,GRMZM2G005732(PRR37/Mal)on chromosome 7 and GRMZM2G004483(CCT)on chromosome 9.These three candidate genes were validated in genome-wide association studies(G WAS)for the trait of total nodal root number(RN).3.These results gave a hint that flowering time played a key role in shaping nodal root number via indirect selection during maize domestication.Our results also supported that more aerial nodal roots and fewer crown roots might be favored in temperate maize,and this root architecture might efficiently improve root-lodging resistance and the ability to take up deep water and nitrogen under dense planting.4.A major QTL controlled maize stiff stalk referred as Stiff1(Stif1)was identified in a recombination inbred line(RIL)population originated from the crosses between a typical stiff inbred line B73 and a non-stiff inbred line Kill,which explained 23.8%and 15.6%of phenotypic variation of stalk bending strength(BS)and rind penetrometer resistance(RPR)respectively.5.Through fine-mapping,Stif1 was narrowed to a 52.9-kb segment containing two genes.The upstream gene encoded an aluminum-activated malate transporter related to stomatal conductance.So the downstream gene containing two exons and one intron was the prior candidate gene of Stif1,which encoded an F-BOX ubiquitination protein with 436 amino acid residues.A 27.2-kb transposon insertion suppressing the expression of Stif1 was found in the promoter region,which was associated with maize stiff stalk.In situ hybridization analysis showed that more transcripts of Stif1 were accumulated within vascular bundles in the stalk of plant without the insertion compared to the plant with the insertion,which was further validated by LUC transient assay.This insertion led to a low content of cellulose and lignin of sclerenchyma cell wall of stalk,then gave rise to a soft stalk and even stalk lodging.6.Phylogenetics analysis indicated that the 27.2-kb transposon insertion in the Stif1 may be singly originated from lowland tropical yellow inbred line CML52.The nucleotide diversity of the promoter of Stif1 was extremely low in the lines with the large insertion.HKA neutral test significantly rejected the neutral null model in CDS regions of Stif1.These results indicated that Stif1 was strongly selected during the improvement of maize.