Analysis of QTLs and Candidate Genes for Tassel Symptoms in Maize Infected with Sporisorium reilianum

Abstract

Heat smut is a fungal soil-borne disease caused by Sporisorium reilianum, and affects the development of male and female tassels. Our previous research found that the tassel symptoms in maize infected with Sporisorium reilianum significantly differed in inbred lines with Sipingtou blood, and exhibited stable heredity over time at multiple locations. In this study, cytological analysis demonstrated that the cellular organization structures of three typical inbred lines (Huangzao4, Jing7, and Chang7-2) showed significant discrepancies at the VT stage. QTLs that control the different symptoms of maize tassels infected with Sporisorium reilianum were located in two F₂ populations, which were constructed using three typical inbred lines. The BSA (bulked segregation analysis) method was used to construct mixed gene pools based on typical tassel symptoms. The QTLs of different symptoms of maize tassels infected with Sporisorium reilianum were detected with 869 SSR markers covering the whole maize genome. The mixed gene pools were screened with polymorphic markers between the parents. Additional SSR markers were added near the above marker to detect genotypes in partially single plants in F₂ populations. The QTL controlling tassel symptoms in the Huangzao4 and Jing7 lines was located on the bin 1.06 region, between the markers of umc1590 and bnlg1598, and explained 21.12% of the phenotypic variation with an additive effect of 0.6524. The QTL controlling the tassel symptoms of the Jing7 and Chang7-2 lines was located on the bin 2.07 region, between the markers of umc1042 and bnlg1335, and explained 11.26% phenotypic variation with an additive effect of 0.4355. Two candidate genes (ZmABP2 and Zm00001D006403) were identified by a conjoint analysis of label-free quantification proteome sequencings.

Keywords: QTL; Sporisorium reilianum; bulked segregation analysis; maize; tassel symptoms.

Figure 1

Cell structure analysis of three typical maize inbred lines and the resistant inbred line Mo17 at different growth stages. (a–d) Cells from Huangzao4 growth points or tassels after inoculation at the VE, V8, V12, and VT stages, respectively. (e–h) Cells from Jing7 growth points or tassels after inoculation at the VE, V8, V12, and VT stages, respectively. (i–l) Cells from Chang7-2 growth points or tassels after inoculation at the VE, V8, V12, and VT stages, respectively. (m–p) Cells from Mo17 growth points or tassels after inoculation at the VE, V8, V12, and VT stages, respectively. The ”-”scale is 1 μm. The red arrow indicates cell autophagy; the green arrow indicates plasmolysis.

Figure 2

Three types of tassel symptoms in typical maize inbred lines infected with Sporisorium reilianum: (a) Huangzao4 normal plant without Sporisorium reilianum; (b) Huangzao4 plant infected with Sporisorium reilianum; (c) Class A with typical symptoms of the tassel of Huangzao4; (d) Jing7 normal plant without Sporisorium reilianum; (e) Jing7 plant infected with Sporisorium reilianum; (f) Class B with typical symptoms of the tassel of Jing7; (g) Chang7-2 normal plant without Sporisorium reilianum; (h) Chang7-2 plant infected with Sporisorium reilianum; and (i) Class C with typical symptoms of the tassel of Chang7-2.

Figure 3

Different tassel symptoms of plants infected with Sporisorium reilianum in F₂ populations: (a) typical Huangzao4 plants in Huangzao4×Jing7 F₂ population; (b) typical Jing7 plants in the Huangzao4×Jing7 F₂ population; (c) typical Chang7-2 plants in the Jing7×Chang7-2 F₂ population.

Figure 4

SSR marker screening among parents. Electrophoresis lanes: M is Marker pBR322 DNA/MspⅠ; the samples are from Huangzao4, Jing7, and Chang7-2, respectively; 1, 2, and 3 are bnlg1083; 4, 5, and 6 are bnlg1614; 7, 8, and 9 are bnlg1953; 10, 11, and 12 are umc1169; 13, 14, and 15 are bnlg1458; 16, 17, and 18 are umc1321; 19, 20, and 21 are umc1073; 22, 23, and 24 are umc1114; 25, 26, and 27 are phi078; 28, 29, and 30 are umc1944; 31, 32, and 33 are umc1095.

Figure 5

Screening of SSR markers between extreme phenotype DNA pools. Electrophoresis lanes: M is Marker pBR322 DNA/MspI; (a) molecular marker of umc1601 and 1, 2, 3, and 4 are Huangzao4, Jing7, Huangzao4 types of the mixed gene pool, and Jing7 types of the mixed gene pool, respectively; (b) molecular marker of umc1590 and 1, 2, 3, and 4 are Huangzao4, Jing7, Huangzao4 types of the mixed gene pool, and Jing7 types of the mixed gene pool; (c) molecular marker of umc1754 and 1, 2, 3, and 4 are Huangzao4, Jing7, Huangzao4 types of the mixed gene pool, and Jing7 types of the mixed gene pool; (d) molecular markers of bnlg1914 and 1, 2, 3, and 4 are Jing7, Chang7-2, Jing7 types of the mixed gene pool, and Chang7-2 types of the mixed gene pool; and (e) molecular marker of bnlg1335 and 1, 2, 3, 4 are Jing7, Chang7-2, Jing7 types of the mixed gene pool, and Chang7-2 types of the mixed gene pool.

Figure 6

QTL results of tassel symptoms of maize infected with Sporisorium reilianum in two F₂ populations. (a) QTL of tassel symptoms of maize infected with Sporisorium reilianum between Huangzao4 and Jing7. (b) QTL of tassel symptoms of maize infected with Sporisorium reilianum between Jing7 and Chang7-2.

Figure 7

Bioinformatics analysis of the candidate gene ZmABP2. (a) Gene prediction of ZmABP2 gene (TSS: Transcription start site; CDSf: Start exon; CDSI: Terminal exon). (b) Tertiary structure of ZmABP2 protein. (c) Phylogenetic analysis of ZmABP2 protein.

Figure 8

Bioinformatics analysis of the candidate gene Zm00001d006403. (a) Gene prediction of Zm00001d006403 gene (TSS: Transcription start site; CDSf: Start exon; CDSI: Terminal exon). (b) Tertiary structure of Zm00001d006403 protein. (c) Phylogenetic analysis of Zm00001d006403 protein.