Crops ›› 2022, Vol. 38 ›› Issue (5): 34-41.doi: 10.16035/j.issn.1001-7283.2022.05.005

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Purity Identification of SSR Molecular Markers for Main Maize Varieties in Jilin Province

Duan Mengran1,2(), Liu Fengze3(), Ge Jianrong1, Yi Hongmei1, Yang Hongming4, Gao Yuqian4, Yue Pengwu4, Ma Wenyu4, Ban Xiuli2,4(), Wang Fengge1()   

  1. 1Beijing Key Laboratory of Maize DNA Fingerprinting and Molecular Breeding, Maize Research Institute, Beijing Academy of Agriculture & Forestry Sciences, Beijing 100097, China
    2College of Agriculture, Jilin Agriculture University, Changchun 130118, Jilin, China
    3National Agriculture Technology Extension Service Center, Beijing 100125, China
    4Seed Management Station of Jilin Province, Changchun 130022, Jilin, China
  • Received:2022-03-11 Revised:2022-08-26 Online:2022-10-15 Published:2022-10-19

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Abstract:

A scientific and efficient purity identification system has been constructed for the main varieties of maize in Jilin province. A total of 50 maize varieties mainly promoted by Jilin province were used as materials. Nine pairs of primers were selected from 40 pairs of SSR primers of GB/T 39914-2021 for the purity identification of the main maize varieties in Jilin. The heterozygosity of the nine pairs of primers were between 82% and 100%, with an average heterozygosity of 91.5%, and the PIC values were between 0.44-0.75, with an average PIC value of 0.63. The set of primers had high polymorphism, strong stability, high resolution, and few non-specific amplification fragments. The nine-fold amplification system was optimized to form a variety purity identification system for main maize varieties in Jilin. The nine pairs of primers were used to identify the purity of the six main varieties in Jilin and could accurately detect the typical strains, inbred seedlings and heterotypic strains of the varieties. A total of ten inbred seedlings and seven heterotypic strains were detected. The highest purity the six samples was 98%, and the lowest purity was 96%. The correlation coefficient between the identification results and the field identification results was 0.766, the correlation between the identification results was high and reliable.

Key words: Maize, Jilin province, Purity, SSR

Table 1

The informations of 50 samples ×104 hm2"

编号
Number		 样品
Sample		 种植面积
Planting area		 编号
Number		 样品
Sample		 种植面积
Planting area		 编号
Number		 样品
Sample		 种植面积
Planting area
1 京科968 22.16 18 九单318 3.01 35 翔玉588 1.70
2 良玉99号 13.54 19 富民985 3.00 36 科泰925 1.69
3 翔玉998 12.01 20 裕丰303 2.99 37 中地9988 1.65
4 天育108 11.81 21 翔玉368 2.95 38 吉东21 1.65
5 先玉335 10.42 22 辽科38 2.74 39 京科665 1.61
6 翔玉211 8.11 23 平安169 2.55 40 九单918 1.58
7 天农九 7.73 24 平安1509 2.38 41 惠民207 1.57
8 优迪919 6.18 25 优旗698 2.06 42 通玉9585 1.57
9 瑞普909 5.77 26 翔玉198 2.05 43 金辉98 1.52
10 杜育311 4.86 27 富民58 2.03 44 军丰6 1.50
11 优旗318 4.16 28 军育288 2.01 45 宏信808 1.39
12 翔玉329 4.09 29 德禹1011 1.91 46 吉农大889 1.37
13 通单258 4.01 30 银河126 1.85 47 MC278 1.33
14 吉农玉719 3.31 31 宏兴528 1.84 48 吉农大935 1.33
15 宏硕313 3.13 32 吉单56 1.77 49 杜育618 1.31
16 优旗511 3.12 33 金园5 1.75 50 远科107 1.29
17 远科1 3.05 34 富民105 1.74

Table 2

SSR primer polymorphism analysis of 50 main maize varieties in Jilin province"

引物
Primer		 基因型数
Genotype number		 等位基因数
Allele number		 等位基因频率
Allele frequency		 基因多样性
Gene diversity		 杂合度
Heterozygosity (%)		 PIC
P01 11 9 0.71 0.48 48 0.46
P02 8 6 0.57 0.59 10 0.53
P03 14 10 0.40 0.71 96 0.66
P04 21 15 0.47 0.74 66 0.72
P05 17 11 0.39 0.76 96 0.73
P06 10 6 0.45 0.67 80 0.62
P07 5 5 0.88 0.22 12 0.21
P08 9 7 0.48 0.61 94 0.55
P09 15 10 0.47 0.71 64 0.68
P10 15 8 0.35 0.74 96 0.69
P11 18 13 0.38 0.78 94 0.75
P12 14 11 0.66 0.54 54 0.53
P13 15 9 0.38 0.74 82 0.70
P14 9 5 0.50 0.66 84 0.60
P15 13 7 0.62 0.59 50 0.56
P16 7 6 0.74 0.43 48 0.41
P17 5 3 0.52 0.61 86 0.55
P18 6 4 0.79 0.36 32 0.33
P19 6 6 0.64 0.52 70 0.47
P20 11 6 0.41 0.67 72 0.61
P21 5 3 0.58 0.52 72 0.42
P22 24 15 0.43 0.77 62 0.76
P23 12 6 0.47 0.62 86 0.55
P24 9 6 0.67 0.49 54 0.44
P25 10 6 0.49 0.69 82 0.65
P26 5 4 0.61 0.52 74 0.43
P27 7 5 0.65 0.53 60 0.49
P28 5 3 0.69 0.44 48 0.37
P29 12 8 0.34 0.72 60 0.66
P30 5 5 0.53 0.54 88 0.44
P31 9 8 0.48 0.67 92 0.62
P32 9 6 0.81 0.34 24 0.32
P33 10 5 0.44 0.65 88 0.59
P34 5 3 0.60 0.52 70 0.43
P35 11 7 0.43 0.65 88 0.59
P36 9 5 0.66 0.53 52 0.49
P37 12 7 0.36 0.73 86 0.69
P38 4 3 0.66 0.46 48 0.37
P39 11 7 0.51 0.60 78 0.53
P40 11 7 0.42 0.66 78 0.59
平均值Mean 10.35 6.9 0.54 0.59 68.1 0.55

Fig.1

Distribution of heterozygous loci of SSR primer combinations in 50 samples"

Fig.2

The heterozygosities of 50 samples in eight pairs, nine pairs and 40 pairs of primers, respectively"

Fig.3

Electrophoresis results of multiple combinations"

Table 3

Factor level of orthogonal experimental design μL"

水平
Level		 2×Taq Plus
Master Mix		 引物Primer
P03、P05、
P08、P31		 P10、P11、
P17、P25		 P37
1 10.00 0.30 0.25 0.20
2 12.00 0.35 0.20 0.15
3 14.00 0.40 0.15 0.10
4 16.00 0.45 0.10 0.05

Table 4

L16(44) orthogonal experiment table μL"

试验号
Test number		 2×Taq Plus
Master Mix		 引物Primer
P03、P05、
P08、P31		 P10、P11、
P17、P25		 P37
1 10.00 0.30 0.25 0.20
2 14.00 0.40 0.25 0.10
3 16.00 0.45 0.25 0.05
4 12.00 0.35 0.25 0.15
5 12.00 0.45 0.15 0.20
6 16.00 0.40 0.20 0.20
7 14.00 0.35 0.10 0.20
8 10.00 0.45 0.10 0.10
9 16.00 0.30 0.10 0.15
10 10.00 0.40 0.15 0.15
11 12.00 0.40 0.10 0.05
12 12.00 0.30 0.20 0.10
13 14.00 0.30 0.15 0.05
14 14.00 0.45 0.20 0.15
15 16.00 0.35 0.15 0.10
16 10.00 0.35 0.20 0.05

Fig.4

Electrophoresis results after optimizing amplification system"

Table 5

Seed purity identification and primer recommendation of 50 main maize varieties in Jilin province"

编号
Number		 样品
Sample		 候选引物
Candidate primer		 优选引物
Optimized primer		 编号
Number		 样品
Sample		 候选引物
Candidate primer		 优选引物
Optimized primer
1 京科968 P03、P08、P10、P17 P03、P10 26 翔玉198 P03、P08、P10、P17 P03、P10
2 良玉99号9 P03、P08、P10、P11 P03、P10 27 富民58 P03、P08、P10、P25 P03、P10
3 翔玉998 P03、P08、P10、P17 P03、P10 28 军育288 P03、P08、P10、P17 P03、P10
4 天育108 P03、P08、P10、P17 P03、P10 29 德禹101 P03、P08、P10、P17 P03、P10
5 先玉335 P03、P08、P10、P17 P03、P10 30 银河126 P03、P08、P10、P25 P03、P10
6 翔玉211 P03、P08、P10、P17 P03、P10 31 宏兴528 P03、P08、P10、P17 P03、P10
7 天农九 P03、P10、P17、P11 P03、P10 32 吉单56 P03、P08、P10、P17 P03、P10
8 优迪919 P03、P08、P10、P17 P03、P10 33 金园5 P03、P08、P10、P17 P03、P10
9 瑞普909 P03、P08、P10、P17 P03、P10 34 富民105 P03、P08、P10、P11 P03、P10
10 杜育311 P03、P08、P10、P17 P03、P10 35 翔玉588 P03、P08、P10、P17 P03、P10
11 优旗318 P03、P08、P10、P17 P03、P10 36 科泰925 P08、P10、P11、P17 P08、P10
12 翔玉329 P03、P08、P10、P17 P03、P10 37 中地9988 P03、P08、P10、P17 P03、P10
13 通单258 P03、P08、P11、P25 P03、P08 38 吉东21 P03、P08、P10、P17 P03、P10
14 吉农玉719 P03、P08、P10、P17 P03、P10 39 京科665 P03、P08、P10、P17 P03、P10
15 宏硕313 P08、P10、P11、P25 P03、P10 40 九单918 P03、P08、P17、P25 P03、P10
16 优旗511 P03、P08、P10、P17 P03、P10 41 惠民207 P03、P08、P10、P17 P03、P10
17 远科1 P03、P08、P10、P17 P03、P10 42 通玉9585 P03、P08、P17、P25 P03、P08
18 九单318 P03、P10、P17、P25 P03、P10 43 金辉98 P03、P10、P17、P25 P03、P10
19 富民985 P03、P08、P10、P17 P03、P10 44 军丰6 P03、P08、P10、P17 P03、P10
20 裕丰303 P03、P08、P10、P17 P03、P10 45 宏信808 P03、P08、P10、P17 P03、P10
21 翔玉368 P03、P08、P10、P17 P03、P10 46 吉农大889 P03、P08、P10、P17 P03、P10
22 辽科38 P03、P08、P10、P17 P03、P10 47 MC278 P03、P08、P10、P17 P03、P10
23 平安169 P03、P08、P10、P17 P03、P10 48 吉农大935 P03、P08、P10、P17 P03、P10
24 平安1509 P03、P08、P10、P17 P03、P10 49 杜育618 P03、P08、P10、P25 P03、P10
25 优旗698 P03、P08、P10、P17 P03、P10 50 远科107 P03、P08、P10、P17 P03、P10

Table 6

Purity identification results of six samples %"

编号
Number		 田间鉴定Field identification SSR分子标记鉴定SSR molecular marker identification
自交苗Inbred seedling 异型株Dimorphic individual 纯度Purity 自交苗Inbred seedling 异型株Dimorphic individual 纯度Purity
D1 2 1 97 2 1 97
D2 4 0 96 4 0 96
D3 2 2 96 2 1 97
D4 1 1 98 1 1 98
D5 1 1 98 1 2 97
D6 0 2 98 0 2 98
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