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Rice black-streaked dwarf virus is transmitted by small brown planthoppers, which causes maize rough dwarf disease and rice black-streaked dwarf disease. This virus leads to slow growth or death of the host plants. During the coevolutionary arms race between viruses and plants, virus-derived small interfering RNAs (vsiRNAs) challenge the plant's defense response and inhibit host immunity through the RNA silencing system. However, it is currently unknown if rice black-streaked dwarf virus can produce the same siRNAs to mediate the RNA silencing in different infected species. In this study, four small RNA libraries and four degradome libraries were constructed by extracting total RNAs from the leaves of the maize ( Zea mays ) inbred line B73 and japonica rice ( Oryza sativa ) variety Nipponbare exposed to feeding by viruliferous and nonviruliferous small brown planthoppers. We analyzed the characteristics of small RNAs and explored virus-derived siRNAs in small RNA libraries through high-throughput sequencing. On analyzing the characteristics of small RNA, we noted that the size distributions of small RNAs were mainly 24 nt (19.74 to 62.00%), whereas those of vsiRNAs were mostly 21 nt (41.06 to 41.87%) and 22 nt (39.72 to 42.26%). The 5'-terminal nucleotides of vsiRNAs tended to be adenine or uracil. Exploring the distribution of vsiRNA hot spots on the viral genome segments revealed that the frequency of hotspots in B73 was higher than those in Nipponbare. Meanwhile, hotspots in the S9 and S10 virus genome segments were distributed similarly in both hosts. In addition, the target genes of small RNA were explored by degradome sequencing. Analyses of the regulatory pathway of these target genes unveiled that viral infection affected the ribosome-related target genes in maize and the target genes in the metabolism and biosynthesis pathways in rice. Here, 562 and 703 vsiRNAs were separately obtained in maize and rice and 73 vsiRNAs named as coexisting vsiRNAs (co-vsiRNAs) were detected in both hosts. Stem-loop PCR and real-time quantitative PCR confirmed that co-vsiRNA 3.1 and co-vsiRNA 3.5, derived from genome segment S3, simultaneously play a role in maize and rice and inhibited host gene expression. The study revealed that rice black-streaked dwarf virus can produce the same siRNAs in different species and provides a new direction for developing new antiviral strategies., Competing Interests: The author(s) declare no conflict of interest.

Insect herbivory challenges plant survival, and coordination of the interactions between growth, herbivore resistance/tolerance is a key problem faced by plants. Based on field experiments into resistance to the Asian corn borer (ACB, Ostrinia furnacalis), we selected 10 inbred maize lines, of which five were resistant and five were susceptible to ACB. We conducted ACB larval bioassays, analysed defensive chemicals, phytohormones, and relative gene expression using RNA-seq and qPCR as well as agronomic traits, and found resistant lines had weaker inducibility, but were more resistant after ACB attack than susceptible lines. Resistance was related to high levels of major benzoxazinoids, but was not related to induced levels of JA or JA-Ile. Following combination analyses of transcriptome, metabolome and larval performance data, we discovered three benzoxazinoids biosynthesis-related transcription factors, NAC60, WRKY1 and WRKY46. Protoplast transformation analysis suggested that these may regulate maize defence-growth trade-offs by increasing levels of benzoxazinoids, JA and SA but decreasing IAA. Moreover, the resistance/tolerance-growth trade-offs were not observed in the 10 lines, and genotype-specific metabolic and genetic features probably eliminated the trade-offs. This study highlights the possibility of breeding maize varieties simultaneously with improved defences and higher yield under complex field conditions., (© 2022 John Wiley & Sons Ltd.)

Maize rough dwarf disease (MRDD) is caused by a virus and seriously affects maize quality and yield worldwide. MRDD can be most effectively controlled with disease-resistant hybrids of corn. Here, MRDD-resistant (Qi319) and -susceptible (Ye478) parental inbred maize lines and their 314 recombinant inbred lines (RILs) that were derived from a cross between them were evaluated across three environments. A stable resistance QTL, qMrdd2 , was identified and mapped using best linear unbiased prediction (BLUP) values to a 0.55-Mb region between the markers MK807 and MK811 on chromosome 2 (B73 RefGen_v3) and was found to explain 8.6 to 11.0% of the total phenotypic variance in MRDD resistance. We validated the effect of qMrdd2 using a chromosome segment substitution line (CSSL) that was derived from a cross between maize inbred Qi319 as the MRDD resistance donor and Ye478 as the recipient. Disease severity index of the CSSL haplotype II harboring qMrdd2 was significantly lower than that of the susceptible parent Ye478. Subsequently, we fine-mapped qMrdd2 to a 315-kb region flanked by the markers RD81 and RD87, thus testing recombinant-derived progeny using selfed backcrossed families. In this study, we identified a novel QTL for MRDD resistance by combining the RIL and CSSL populations, thus providing important genetic information that can be used for breeding MRDD-resistant varieties of maize.

Background: Natural variations derived from both evolutionary selection and genetic recombination, presume to have important functions to respond to various abiotic stresses, which could be used to improve drought tolerance via genomic selection., Results: In the present study, the NAC-encoding gene of ZmNAC080308 was cloned and sequenced in 199 inbred lines in maize. Phylogenetic analysis showed that ZmNAC080308 is closely clusteredinto the same group with other well-known NAC genes responding to improve drought tolerance. In total, 86 SNPs and 47 InDels were identified in the generic region of ZmNAC080308, 19 of these variations were associated with GY (grain yield) in different environments. Nine variations in the 5'-UTR region of ZmNAC080308 are closely linked, they might regulate the gene expression and respond to improve GY under drought condition via Sp1-mediated transactivation. Two haplotypes (Hap1 and Hap2) identified in the, 5'-UTR region using the nine variations, and Hap2 containing insertion variants, exhibited 15.47 % higher GY under drought stress condition. Further, a functional marker was developed to predict the drought stress tolerance in a US maize inbred line panel. Lines carrying Hap2 exhibited > 10 % higher GY than those carrying Hap1 under drought stress condition. In Arabidopsis, overexpression ZmNAC080308 enhanced drought tolerance., Conclusions: ZmNAC080308 is an important gene responding to drought tolerance, a functional marker is developed for improving maize drought tolerance by selecting this gene.

Background: Starch biosynthesis in endosperm is a key process influencing grain yield and quality in maize. Although a number of starch biosynthetic genes have been well characterized, the mechanisms by which the expression of these genes is regulated, especially in regard to microRNAs (miRNAs), remain largely unclear., Results: Sequence data for small RNAs, degradome, and transcriptome of maize endosperm at 15 and 25 d after pollination (DAP) from inbred lines Mo17 and Ji419, which exhibit distinct starch content and starch granule structure, revealed the mediation of starch biosynthetic pathways by miRNAs. Transcriptome analysis of these two lines indicated that 33 of 40 starch biosynthetic genes were differentially expressed, of which 12 were up-regulated in Ji419 at 15 DAP, one was up-regulated in Ji419 at 25 DAP, 14 were up-regulated in Ji419 at both 15 and 25 DAP, one was down-regulated in Ji419 at 15 DAP, two were down-regulated in Ji419 at 25 DAP, and three were up-regulated in Ji419 at 15 DAP and down-regulated in Ji419 at 25 DAP, compared with Mo17. Through combined analyses of small RNA and degradome sequences, 22 differentially expressed miRNAs were identified, including 14 known and eight previously unknown miRNAs that could target 35 genes. Furthermore, a complex co-expression regulatory network was constructed, in which 19 miRNAs could modulate starch biosynthesis in endosperm by tuning the expression of 19 target genes. Moreover, the potential operation of four miRNA-mediated pathways involving transcription factors, miR169a-NF-YA1-GBSSI/SSIIIa and miR169o-GATA9-SSIIIa/SBEIIb, was validated via analyses of expression pattern, transient transformation assays, and transactivation assays., Conclusion: Our results suggest that miRNAs play a critical role in starch biosynthesis in endosperm, and that miRNA-mediated networks could modulate starch biosynthesis in this tissue. These results have provided important insights into the molecular mechanism of starch biosynthesis in developing maize endosperm.

Key Message: Using combined linkage and association mapping, 26 stable QTL and six stable SNPs were detected across multiple environments for eight ear and grain morphological traits in maize. One QTL, PKS2, might play an important role in maize yield improvement. In the present study, one bi-parental population and an association panel were used to identify quantitative trait loci (QTL) for eight ear and grain morphological traits. A total of 108 QTL related to these traits were detected across four environments using an ultra-high density bin map constructed using recombinant inbred lines (RILs) derived from a cross between Ye478 and Qi319, and 26 QTL were identified in more than two environments. Furthermore, 64 single nucleotide polymorphisms (SNPs) were found to be significantly associated with the eight ear and grain morphological traits (-log 10 (P) > 4) in an association panel of 240 maize inbred lines. Combining the two mapping populations, a total of 17 pleiotropic QTL/SNPs (pQTL/SNPs) were associated with various traits across multiple environments. PKS2, a stable locus influencing kernel shape identified on chromosome 2 in a genome-wide association study (GWAS), was within the QTL confidence interval defined by the RILs. The candidate region harbored a short 13-Kb LD block encompassing four SNPs (SYN11386, PHM14783.16, SYN11392, and SYN11378). In the association panel, 13 lines derived from the hybrid PI78599 possessed the same allele as Qi319 at the PHM14783.16 (GG) locus, with an average value of 0.21 for KS, significantly lower than that of the 34 lines derived from Ye478 that carried a different allele (0.25, P < 0.05). Therefore, further fine mapping of PKS2 will provide valuable information for understanding the genetic components of grain yield and improving molecular marker-assisted selection (MAS) in maize.

Key Message: A QTL qMrdd8 that confers resistance to MRDD was fine mapped into an interval of 347 kb; one SNP and two InDels identified in the interval were significantly associated with resistance to MRDD. Maize rough dwarf disease (MRDD) is highly prevalent in the summer maize-growing areas in China, and leads to significant yield losses in maize (Zea mays L.). In this study, the quantitative trait locus (QTL) qMrdd8, which confers resistance to MRDD, was fine mapped. Initially, qMrdd8 was consistently identified in the interval between the simple sequence repeat markers umc1617 and phi121 in three F 2 sub-populations derived from a cross between the resistant recombinant inbred line NL203 and the susceptible line B73. Subsequently, qMrdd8 was fine mapped into an interval of 347 kb defined by the markers IDRQ2 and IDRQ20 using a recombinant-derived progeny test strategy. Based on single nucleotide polymorphism (SNP) genotypes identified using the MaizeSNP50 BeadChip, a long haplotype including qMrdd8 was identified in four resistant inbred lines. One SNP, the 2549-bp insertion/deletion polymorphism (InDel) InDel25, and the 2761-bp InDel27, which all were significantly associated with resistance to MRDD in a set of 226 maize inbred lines (P < 0.05), were detected within qMrdd8. Furthermore, two candidate genes, CG1 and CG2, were detected in the interval using RNA sequencing (RNA-Seq), and InDel25 was localized within the candidate gene CG1. In conclusion, the fine mapping of qMrdd8 will be helpful in cloning the resistance gene, and the three polymorphic markers identified in this study could be used to improve MRDD resistance via a marker-assisted selection approach.

To analyze the variation in rice black-streaked dwarf virus (RBSDV) in an area with high incidence of maize rough dwarf disease (MRDD), the RBSDV S1 segment in a collection of 100 maize isolates (sample population A100) from Jining, Shandong Province, was sequenced. An additional 21 maize and rice isolates (subpopulation B21) that were sampled from nine other geographic locations in China in 2012 and 2013 were used as a control. A total of 914 nucleotide mutations, including 239 singleton variable and 675 parsimony-informative sites were detected among the segment 1 (S1) sequences from A100. A total of 614 nucleotide mutation sites including 164 singleton variable and 450 parsimony-informative sites were detected among the S1 sequences from B21, while 97.55 % of the parsimony-informative sites from B21 were also detected in A100. The nucleotide sequence diversities of A100 (π = 0.0479) and B21 (π = 0.0396) were significantly different (P = 0.0002) but showed similar trends. Phylogenetic analysis showed that the 121 RBSDV isolates could be classified into two groups based on their S1 sequences, independent of subpopulation, with a combination of host species and locations. A100 and B21 were under the same level of negative and purifying selection, with Ka/Ks ratios of 0.0337 and 0.0369, respectively. The combined RBSDV population, including 121 isolates, was expanding, with negative values for Tajima's D, Fu and Li's D, and Fu and Li's F in both A100 and B21, except Tajima's D in A100. Based on S1, the RBSDV population in China has long-term phytogeographic stability, and there do not appear to be any newly-emerging strains.

Maize rough dwarf disease (MRDD) is a viral infection that results in heavy yield losses in maize worldwide, particularly in the summer maize-growing regions of China. MRDD is caused by the Rice black-streaked dwarf virus (RBSDV). In the present study, analyses of microRNAs (miRNAs), the degradome, and transcriptome sequences were used to elucidate the RBSDV-responsive pathway(s) in maize. Genomic analysis indicated that the expression of three non-conserved and 28 conserved miRNAs, representing 17 known miRNA families and 14 novel miRNAs, were significantly altered in response to RBSDV when maize was inoculated at the V3 (third leaf) stage. A total of 99 target transcripts from 48 genes of 10 known miRNAs were found to be responsive to RBSDV infection. The annotations of these target genes include a SQUAMOSA promoter binding (SPB) protein, a P450 reductase, an oxidoreductase, and a ubiquitin-related gene, among others. Characterization of the entire transcriptome suggested that a total of 28 and 1085 differentially expressed genes (DEGs) were detected at 1.5 and 3.0 d, respectively, after artificial inoculation with RBSDV. The expression patterns of cell wall- and chloroplast-related genes, and disease resistance- and stress-related genes changed significantly in response to RBSDV infection. The negatively regulated genes GRMZM2G069316 and GRMZM2G031169, which are the target genes for miR169i-p5 and miR8155, were identified as a nucleolin and a NAD(P)-binding Rossmann-fold superfamily protein in maize, respectively. The gene ontology term GO:0003824, including GRMZM2G031169 and other 51 DEGs, was designated as responsive to RBSDV., (© The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Waxy maize is prevalently grown in China and other countries due to the excellent characters and economic value. However, its low content of lysine can't meet the nutritional requirements of humans and livestock. In the present study, we introgressed the opaque2 (o2) allele into waxy maize line Zhao OP-6/O2O2 by using marker-assisted selection (MAS) technique and successfully improved the lysine content and quality of waxy maize. Transcript abundance analysis indicated that the wx1 expression levels had no difference between Zhao OP-6/o2o2 and Zhao OP-6/O2O2. However, Zhao OP-6/o2o2 was characterized by a phenotype of hard and vitreous kernels and accumulation of protein bodies at smaller size (one third of that of parents) but in larger numbers. Biochemical analyses showed that Zhao OP-6/o2o2 had 16.7% less free amino acids than Zhao OP-6/O2O2, especially those derived from glycolytic intermediates, but its content of lysine was increased by 51.6% (0.47% vs. 0.31%). The content of amylopectin was 98.5% in Zhao OP-6/o2o2, significantly higher than that in Zhao OP-6/O2O2 (97.7%). Proteomic analyses indicated that o2 introgression not only decreased the accumulation of various zein proteins except for 27-kDa γ-zein, but also affected other endosperm proteins related to amino acid biosynthesis, starch-protein balance, stress response and signal transduction. This study gives us an intriguing insight into the metabolism changes in endosperm of waxy maize introgressed with opaque2.

Background: Plant architecture attributes, such as plant height, ear height, and internode number, have played an important role in the historical increases in grain yield, lodging resistance, and biomass in maize (Zea mays L.). Analyzing the genetic basis of variation in plant architecture using high density QTL mapping will be of benefit for the breeding of maize for many traits. However, the low density of molecular markers in existing genetic maps has limited the efficiency and accuracy of QTL mapping. Genotyping by sequencing (GBS) is an improved strategy for addressing a complex genome via next-generation sequencing technology. GBS has been a powerful tool for SNP discovery and high-density genetic map construction. The creation of ultra-high density genetic maps using large populations of advanced recombinant inbred lines (RILs) is an efficient way to identify QTL for complex agronomic traits., Results: A set of 314 RILs derived from inbreds Ye478 and Qi319 were generated and subjected to GBS. A total of 137,699,000 reads with an average of 357,376 reads per individual RIL were generated, which is equivalent to approximately 0.07-fold coverage of the maize B73 RefGen&#95;V3 genome for each individual RIL. A high-density genetic map was constructed using 4183 bin markers (100-Kb intervals with no recombination events). The total genetic distance covered by the linkage map was 1545.65 cM and the average distance between adjacent markers was 0.37 cM with a physical distance of about 0.51 Mb. Our results demonstrated a relatively high degree of collinearity between the genetic map and the B73 reference genome. The quality and accuracy of the bin map for QTL detection was verified by the mapping of a known gene, pericarp color 1 (P1), which controls the color of the cob, with a high LOD value of 80.78 on chromosome 1. Using this high-density bin map, 35 QTL affecting plant architecture, including 14 for plant height, 14 for ear height, and seven for internode number were detected across three environments. Interestingly, pQTL10, which influences all three of these traits, was stably detected in three environments on chromosome 10 within an interval of 14.6 Mb. Two MYB transcription factor genes, GRMZM2G325907 and GRMZM2G108892, which might regulate plant cell wall metabolism are the candidate genes for qPH10., Conclusions: Here, an ultra-high density accurate linkage map for a set of maize RILs was constructed using a GBS strategy. This map will facilitate identification of genes and exploration of QTL for plant architecture in maize. It will also be helpful for further research into the mechanisms that control plant architecture while also providing a basis for marker-assisted selection.

The recessive opaque-2 mutant gene (o2) reduces α-zeins accumulation in maize endosperm, changes the amino acid composition of maize kernels, induces an opaque endosperm, and increases the lysine content of kernels. The quality protein maize (QPM) inbred line CA339 (o2o2) and an elite normal inbred line liao2345 (O2O2) were used to construct o2 near-isogenic lines (NILs) by marker-assisted selection (MAS) using the co-dominant SSR marker phi057. Two specific o2 NILs were constructed, named liao2345/o2-1 and liao2345/o2-2. However, the kernel phenotypes of the two o2 NILs were different from each other. liao2345/o2-1 had the wild-type vitreous endosperm, which is similar to its recurrent parent liao2345, while the endosperm of liao2345/o2-2 was opaque, identical to typical o2 mutant individuals. In comparison to their recurrent parent liao2345, the lysine concentration of liao2345/o2-1 was similar and the lysine concentration in liao2345/o2-2 was doubled. SDS-PAGE analysis indicated that liao2345/o2-1 had the same zeins ratio as liao2345, whereas the zeins concentration of liao2345/o2-2 was markedly lower. Sequence and transcript abundance analyses indicated that the CDS of two o2 NILs are derived from CA339, but they have different promoters. The O2 transcript of liao2345/o2-2 is largely inhibited because of an rbg transposable element inserted between the TATA box and initiator codon of liao2345/o2-2. We concluded that different crossing-over patterns during the process of o2 NIL construction resulted in the different kernel phenotypes of the two o2 NILs. We surmise that the reversion of liao2345/o2-1 to wild type was due to the recombination with the wild type liao2345 promoter during introgression and backcrossing. The o2 mutant gene of donor (CA339) is a null mutant because of low O2 expression. However, its CDS probably encodes a protein with normal function which can maintain the normal accumulation of zeins in maize endosperm.

Background: Kernel weight, controlled by quantitative trait loci (QTL), is an important component of grain yield in maize. Cytokinins (CKs) participate in determining grain morphology and final grain yield in crops. ZmIPT2, which is expressed mainly in the basal transfer cell layer, endosperm, and embryo during maize kernel development, encodes an isopentenyl transferase (IPT) that is involved in CK biosynthesis., Results: The coding region of ZmIPT2 was sequenced across a panel of 175 maize inbred lines that are currently used in Chinese maize breeding programs. Only 16 single nucleotide polymorphisms (SNPs) and seven haplotypes were detected among these inbred lines. Nucleotide diversity (π) within the ZmIPT2 window and coding region were 0.347 and 0.0047, respectively, and they were significantly lower than the mean nucleotide diversity value of 0.372 for maize Chromosome 2 (P < 0.01). Association mapping revealed that a single nucleotide change from cytosine (C) to thymine (T) in the ZmIPT2 coding region, which converted a proline residue into a serine residue, was significantly associated with hundred kernel weight (HKW) in three environments (P <0.05), and explained 4.76% of the total phenotypic variation. In vitro characterization suggests that the dimethylallyl diphospate (DMAPP) IPT activity of ZmIPT2-T is higher than that of ZmIPT2-C, as the amounts of adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) consumed by ZmIPT2-T were 5.48-, 2.70-, and 1.87-fold, respectively, greater than those consumed by ZmIPT2-C. The effects of artificial selection on the ZmIPT2 coding region were evaluated using Tajima's D tests across six subgroups of Chinese maize germplasm, with the most frequent favorable allele identified in subgroup PB (Partner B)., Conclusions: These results showed that ZmIPT2, which is associated with kernel weight, was subjected to artificial selection during the maize breeding process. ZmIPT2-T had higher IPT activity than ZmIPT2-C, and this favorable allele for kernel weight could be used in molecular marker-assisted selection for improvement of grain yield components in Chinese maize breeding programs.

Maize rough dwarf disease (MRDD, a viral disease) results in significant grain yield losses, while genetic basis of which is largely unknown. Based on comparative genomics, eukaryotic translation initiation factor 4E (eIF4E) was considered as a candidate gene for MRDD resistance, validation of which will help to understand the possible genetic mechanism of this disease. ZmeIF4E (orthologs of eIF4E gene in maize) encodes a protein of 218 amino acids, harboring five exons and no variation in the cDNA sequence is identified between the resistant inbred line, X178 and susceptible one, Ye478. ZmeIF4E expression was different in the two lines plants treated with three plant hormones, ethylene, salicylic acid, and jasmonates at V3 developmental stage, suggesting that ZmeIF4E is more likely to be involved in the regulation of defense gene expression and induction of local and systemic resistance. Moreover, four cis-acting elements related to plant defense responses, including DOFCOREZM, EECCRCAH1, GT1GAMSCAM4, and GT1CONSENSUS were detected in ZmeIF4E promoter for harboring sequence variation in the two lines. Association analysis with 163 inbred lines revealed that one SNP in EECCRCAH1 is significantly associated with CSI of MRDD in two environments, which explained 3.33 and 9.04 % of phenotypic variation, respectively. Meanwhile, one SNP in GT-1 motif was found to affect MRDD resistance only in one of the two environments, which explained 5.17 % of phenotypic variation. Collectively, regulatory motifs respectively harboring the two significant SNPs in ZmeIF4E promoter could be involved in the defense process of maize after viral infection. These results contribute to understand maize defense mechanisms against maize rough dwarf virus.

The major quantitative trait locus (QTL) qHS2.09 plays an important role in resistance to head smut during maize breeding and production. In this study, a near-isogenic line (NIL), L34, which harbors the major QTL qHS2.09 in bin 2.09, was developed using a resistant donor 'Mo17' in a susceptible genetic background 'Huangzao4'. Using 18,683 genome-wide polymorphic loci, this major QTL was finely mapped into an interval of ≈1.10 Mb, flanked by single nucleotide polymorphism (SNP) markers PZE-102187307 and PZE-102188421. Moreover, the favorable allele from 'Mo17' for SNP PZE-102187611 in this interval that was most significantly associated with resistance to head smut (P = 1.88 E-10) and accounted for 39.7 to 44.4% of the phenotypic variance in an association panel consisting of 80 inbred lines. With combined linkage and association mapping, this major QTL was finally located between SNP PZE-102187486 and PZE-102188421 with an interval of ≈1.00 Mb. Based on the pedigrees of 'Mo17' and its derivatives widely used in temperate maize breeding programs, the favorable haplotype from 'Mo17' is shown to be the main source of resistance to head smut in these lines. Therefore, the SNPs closely linked to the major QTL qHS2.09, detected in both linkage and association mapping, and could be useful for marker-assisted selection in maize breeding programs.

The recessive mutant allele of the opaque2 gene (o2) alters the endosperm protein pattern and increases the kernel lysine content of maize (Zea mays L.). In this study, sequencing results showed that the o2 mutant was successfully introgressed into 12 elite normal maize inbred lines by marker assisted selection (MAS). The average genetic similarity between these normal inbred lines and their o2 near-isogenic lines (NILs) was more than 95%. Kernel lysine content increased significantly in most of o2 NILs lines relative to normal elite inbreds, but remained unchanged in the genetic backgrounds Dan598o2 and Liao2345o2. Moreover, the kernel characteristics of these two o2 NILs did not differ from the other inbred lines. The results of lysine content analysis in the F1 hybrids between Liao2345o2 and Dan598o2 and other o2 NILs demonstrated that gene(s) other than opaque2 may control kernel lysine content in these two o2 NILs. The results of zein analysis showed that 22-kD α-zein synthesis was reduced or absent, and the 19-kD α-zein synthesis was greatly reduced compared with the recurrent parents in most o2 NILs except for Dan598o2 and Liao2345o2. Our results indicate that gene(s) other than opaque2 may play more important roles in zein synthesis and kernel lysine content in some maize genetic backgrounds.

Single nucleotide polymorphism (SNP) is a common form of genetic variation and popularly exists in maize genome. An Illumina GoldenGate assay with 1 536 SNP markers was used to genotype maize inbred lines and identified the functional genetic variations underlying drought tolerance by association analysis. Across 80 lines, 1 006 polymorphic SNPs (65.5% of the total) in the assay with good call quality were used to estimate the pattern of genetic diversity, population structure, and familial relatedness. The analysis showed the best number of fixed subgroups was six, which was consistent with their original sources and results using only simple sequence repeat markers. Pairwise linkage disequilibrium (LD) and association mapping with phenotypic traits investigated under water-stressed and well-watered regimes showed rapid LD decline within 100-500 kb along the physical distance of each chromosome, and that 29 SNPs were associated with at least two phenotypic traits in one or more environments, which were related to drought-tolerant or drought-responsive genes. These drought-tolerant SNPs could be converted into functional markers and then used for maize improvement by marker-assisted selection., (© 2011 Institute of Botany, Chinese Academy of Sciences.)

Plant invertases catalyze the conversion of sucrose to glucose and fructose, which are distinct signals of widely varied stress-tolerance processes, including the biosynthesis and detection of hormones under water deficit. In the invertase gene family, the candidate gene ivr2 encoding plant acid-soluble invertase plays a vital role in drought tolerance. In this study, a putative genomic sequence of ivr2 including three exons and two introns was acquired and genetically analyzed using bioinformatics and statistics, based on a partial ivr2 gene sequence of the GenBank library. The ivr2 genomic sequence data from 106 maize inbred lines were obtained using five nested primer pairs. Further analysis showed that the detected polymorphic sites were mainly located in exon-1, intron-1 and exon-2 regions; High linkage disequilibrium level and low nucleotide diversity were identified at this ivr2 locus. Association mapping combined the genotypic and phenotypic data, and a total of 48 associations showed high contributions to the variations in grain yield and its components under well-watered and water-stressed conditions over 2 years of experiments. This suggested that functional polymorphisms in ivr2 were possibly associated with maize drought tolerance. This provides reference information for efficient marker-assisted selection of superior alleles in drought tolerance breeding programs.

Background: The harvest index for many crops can be improved through introduction of dwarf stature to increase lodging resistance, combined with early maturity. The inbred line Shen5003 has been widely used in maize breeding in China as a key donor line for the dwarf trait. Also, one major quantitative trait locus (QTL) controlling plant height has been identified in bin 5.05-5.06, across several maize bi-parental populations. With the progress of publicly available maize genome sequence, the objective of this work was to identify the candidate genes that affect plant height among Chinese maize inbred lines with genome wide association studies (GWAS)., Methods and Findings: A total of 284 maize inbred lines were genotyped using over 55,000 evenly spaced SNPs, from which a set of 41,101 SNPs were filtered with stringent quality control for further data analysis. With the population structure controlled in a mixed linear model (MLM) implemented with the software TASSEL, we carried out a genome-wide association study (GWAS) for plant height. A total of 204 SNPs (P≤0.0001) and 105 genomic loci harboring coding regions were identified. Four loci containing genes associated with gibberellin (GA), auxin, and epigenetic pathways may be involved in natural variation that led to a dwarf phenotype in elite maize inbred lines. Among them, a favorable allele for dwarfing on chromosome 5 (SNP PZE-105115518) was also identified in six Shen5003 derivatives., Conclusions: The fact that a large number of previously identified dwarf genes are missing from our study highlights the discovery of the consistently significant association of the gene harboring the SNP PZE-105115518 with plant height (P=8.91e-10) and its confirmation in the Shen5003 introgression lines. Results from this study suggest that, in the maize breeding schema in China, specific alleles were selected, that have played important roles in maize production., (© 2011 Weng et al.)