Crops ›› 2020, Vol. 36 ›› Issue (4): 99-106.doi: 10.16035/j.issn.1001-7283.2020.04.014

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The Development of waxy Gene Function Marker and Its Application in Waxy Maize Breeding

Yuan Wenya1(), Zhao Xiaolei1(), Zhou Xumei2, Wang Lei3, Peng Bo1(), Wang Yi1()   

  1. 1Tianjin Crop Research Institute/Tianjin Key Laboratory of Crop Genetics and Breeding, Tianjin 300384, China
    2Dandong Academy of Agricultural Sciences, Fengcheng 118109, Liaoning, China
    3Handan Academy of Agricultural Sciences, Handan 056001, Hebei, China
  • Received:2019-12-05 Revised:2020-01-09 Online:2020-08-15 Published:2020-08-11
  • Contact: Peng Bo,Wang Yi E-mail:ywenya0916@163.com;xiaoleizhao1981@163.com;snbopeng@163.com;wangyishengji@163.com

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Abstract:

To accelerate the improvement of waxy maize germplasm resources by molecular marker-assisted selection, waxy maize inbred line Yun 539 and ordinary maize inbred line Chang 7-2 were used. Through DNA sequence analysis, it was found that the wx gene mutation of Yun 539 belongs to wx-D7 mutation. The waxy gene function marker FMD7 was developed based on sequence difference between the two inbred lines and it was identified through phenotypic verification and universal verification. By using the marker FMD7 for molecular marker-assisted selection combined with yield, plant type, and quality trait analysis, six excellent Chang 7-2 improved lines carrying wx gene were identified. The marker FMD7 developed in this study for molecular selection can speed up the process of backcross breeding, improve the breeding efficiency, and provide a material basis for breeding high-yielding waxy maize hybrids. At the same time, it provides a theoretical reference for molecular marker-assisted selection of traits controlled by single gene or a few major genes.

Key words: Maize, Gene, FMD7, Molecular marker-assisted selection

Table 1

Primers for segmental amplification of the waxy gene"

编号
Number		 引物名称
Primer name		 5′-3′引物序列
5′-3′ primer sequence		 退火温度
Annealing temperature (℃)		 扩增条带长度
Length of amplify band(bp)
A-F Wxe3-7 GTCTTCTTCGTGCTCTTGCC 57.4 907
A-R GATGCCGTGGGACTGGTAG 58.2
B-F Wxe4-9 GTTGACCACCCACTGTTCCT 56.6 880
B-R ATGAGCTCCTCGGCGTAGTA 57.6
C-F Wxe7-10 AACTACCAGTCCCACGGCATCT 62.4 902
C-R CACGTCCTCCACCATCTCCAT 61.7
D-F Wxe9-13 TGCGAGCTCGACAACATCATGCG 70.9 1 344
D-R AGGGCGCGGCCACGTTCTCC 73.0
E-F Wxe11-14 GAGAAGTTCCCAGGCAAGGT 58.1 931
E-R AGCACAAGCAAGCAGCTACA 57.1

Fig.1

The deletion of 30bp in the 7th exon-7th intron of wx allele gene and the 10th exon sequence in Yun 539"

Table 2

Sequences and annealing temperatures of FMD7"

序号
Number		 引物
Primer		 5′-3′序列
5′-3′ sequence		 退火温度
Annealing
temperature (℃)
1 FMD7-F TACCAGTCCCACGGCATCTA 58.6
2 FMD7-R CACCACCACTACCAGACGAG 55.7

Fig.2

Development of wx function marker of waxy gene in Yun 539"

Table 3

Genotype and phenotype identification of F2 populations"

基因型
Genotype		 籽粒数Kernel number X2 P
基因型鉴定
Genotype
identification		 表型鉴定
Phenotype
identification
wxwx 54 51 1.696 0.638
Wxwx 86 86
WxWx 47 46
其他Other 3 7

Table 4

Marker analysis of FMDT and waxy property of tested maize materials"

编号Number 自交系Inbred line Wx/wx FMD7
1 昌7-2 Wx -
2 郑58 Wx -
3 w359 wx +
4 w360 wx +
5 云539 wx +
6 云413 wx +
7 98-1329 wx +
8 w441 wx -
9 w272 wx -
10 F002 wx +

Fig.3

Sequence variation of the major mutant sites in 10 maize inbred lines The black dotted box is the mutation site of type wx-D10 and the red implementation box is the 7th exon region"

Table 5

Analysis of variance for yield traits among 6 Chang 7-2 waxy improved families, donors and recurrent parents"

变异来源
Variation source		 单株产量
Grain yield
per plant		 穗粗
Ear
diameter		 穗长
Ear
length		 穗行数
Row
number		 行粒数
Kernels
per row		 秃尖长
Bare tip
length		 轴粗
Shaft
diameter		 百粒重
100-kernel
weight		 出籽率
Rate of
seed
基因型Genotype 597.43** 0.04 0.75 4.57 45.14* 0.88** 0.11* 2.49** 0.59**
重复Repeat 9.00 2.00E-03 0.09 4.00 9.00 0.00 3.00E-03 0.30 9.00E-03
误差Error 66.14 0.11 3.26 4.00 6.71 0.04 0.02 0.34 0.08

Table 6

Performance of six Chang 7-2 waxy improved families, donors and recurrent parents"

编号
Number		 单株产量
Grain yield
per plant (g)		 穗粗
Ear diameter
(cm)		 穗长
Ear length
(cm)		 穗行数
Row
number		 行粒数
Kernels
per row		 秃尖长
Bare tip
length (cm)		 轴粗
Cob diameter
(cm)		 百粒重
100-kernel
weight (g)		 出籽率
Rate of
seed (%)
云539 Yun 539 60.0c 4.5 13.4 14.0 22d 1.5a 2.5ab 22.7bc 77.0b
昌7-2 Chang 7-2 88.0b 4.1 12.0 16.0 31bc 0.0b 2.2bc 22.8bc 88.0a
11 108.5ab 4.4 13.2 16.0 38ab 0.2b 2.4b 23.8ab 83.6ab
18 97.1b 4.3 13.1 16.0 27cd 1.5a 2.8a 24.8a 69.2c
24 119.1a 4.3 14.0 16.0 34ab 0.0b 2.5ab 25.2a 81.3ab
38 102.3ab 4.4 12.5 18.0 28bcd 0.0b 2.3bc 24.6a 79.9b
39 96.4b 4.3 13.5 18.0 30bc 0.0b 2.3bc 22.1c 80.9b
64 95.5b 4.1 13.1 14.0 30bc 0.5b 2.0c 24.0b 81.8ab

Table 7

Variance analysis of plant type traits in six Chang 7-2 waxy improved families, donors and recurrent parents"

变异来源Variation source 株高Plant height 穗位高Ear height 雄穗长Tassel length 雄穗分枝数Tassel branch number 叶夹角Leaf angle
基因型Genotype 243.68** 157.85** 12.25 27.86 28.85**
重复Repeat 6.25 0.56 2.25 0.25 14.06
误差Error 19.39 5.71 3.96 11.68 2.78

Table 8

Plant type traits of six Chang 7-2 waxy improved families, donors and recurrent parents"

编号
Number		 株高
Plant height (cm)		 穗位高
Ear height (cm)		 雄穗长
Tassel length (cm)		 雄穗分枝数
Tassel branch number		 叶夹角
Leaf angle (°)
云539 Yun 539 155.4b 62.2b 29.3 9.3 29.0a
昌7-2 Chang 7-2 185.4a 87.3a 22.2 19.3 21.0cd
11 184.0a 88.5a 23.3 18.2 18.5d
18 189.2a 88.2a 23.3 21.3 19.5cd
24 186.3a 87.4a 22.1 19.4 17.0d
38 188.3a 87.3a 22.3 19.2 24.5b
39 185.3a 87.5a 23.4 17.2 23.0cd
64 183.1a 83.0a 21.1 20.1 21.0cd
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