Creation and evaluation for corn germplasm of double recessive sweet-waxy genotype
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摘要:目的
创制甜糯基因双隐性玉米自交系，了解甜糯基因双隐性对品质性状的影响，为甜糯玉米育种提供参考和依据。
方法以1份甜玉米Zea mays ssp. saccharata骨干自交系(M01)和3份糯玉米Z. mays var. ceratina自交系(L33、L35和L38)为材料，用与糯玉米隐性基因wx紧密连锁的分子标记辅助筛选，F2代得到甜糯双隐性玉米材料，连续自交，F4代得到11个纯合甜糯双隐性玉米自交系，检测其可溶性糖含量、淀粉含量及乳熟期籽粒果皮厚度，并与对应的甜、糯玉米亲本进行比较分析。
结果与wx基因紧密连锁的引物phi061在甜、糯玉米亲本间扩增出清晰且呈共显性的条带，可在F2代作为糯质基因的前景选择标记。11份甜糯双隐性自交系材料的可溶性糖质量分数平均为12.27%，比甜玉米亲本高2.55%，远高于糯玉米亲本；淀粉质量分数平均为19.73%，比甜玉米亲本低3.77%，远低于糯玉米亲本；乳熟期籽粒冠部和背胚部果皮厚度均介于对应的杂交亲本之间。
结论通过分子标记辅助选择可快速创制甜糯纯合双隐性玉米材料，其可溶性糖含量高于甜质亲本，淀粉含量低于甜质亲本，果皮厚度介于双亲之间。
Abstract:ObjectiveTo create double recessive sweet-waxy corn inbred lines, identify the effects of double recessive sweet-waxy genes on quality traits, and provide a reference and basis for sweet-waxy corn breeding.
MethodThe double recessive sweet-waxy corn materials with wxwxsh2sh2 genotype were screened by molecular marker that linked with waxy corn recessive gene wx in F2 populations obtained from one elite sweet corn line (M01) and three waxy corn lines (L33, L35 and L38). Then these materials were self-pollinated continuously, and 11 pure double recessive sweet-waxy corn inbred lines with wxwxsh2sh2 genotype were obtained in F4 populations. Their soluble sugar content, total starch content and pericarp thickness were measured and compared with the corresponding sweet corn parent and waxy corn parent.
ResultThe primer phi061, which was tightly linked to wx gene, amplified clear and codominant bands between sweet corn parent and waxy corn parent and was selected as the foreground selection marker of waxy gene. The average content of soluble sugar was 12.27% for double recessive sweet-waxy corn inbred lines, which was 2.55% higher than that sweet corn parent and far higher than that waxy corn parent. The average total starch content was 19.73%, which was 3.77% lower than that sweet corn parent, and far lower than that waxy corn parent. The pericarp thicknesses of crown and abgerminal regions of double recessive maize lines were in the interval of their corresponding parents.
ConclusionIt is a feasible way to create rapidly the double recessive sweet-waxy corn lines by molecular marker assisted selection. Their average content of soluble sugar is higher than that of sweet corn parent, the average content of starch is lower than that of sweet parent, and the average pericarp thickness is between their parents.
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Keywords:
- sweet-waxy corn /
- molecular marker-assisted selection /
- soluble sugar /
- starch /
- pericarp thickness
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图 1 引物phi022、phi027、phi061在甜玉米自交系和糯玉米自交系之间的多态性筛选
Figure 1. The polymorphism screenings between sweet corn line (M01) and wax corn lines (L33, L35 and L38) by phi022, phi027 and phi061 primers
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图 2 利用引物phi061鉴定L35×M01得到的F2群体中的双隐性植株
1~21为以L35×M01得到的F2群体全基因组DNA为模板的PCR扩增产物，其中1、8、11和12为双隐性纯合单株；M01：甜玉米亲本，L35：糯玉米亲本
Figure 2. The identifications of double recessive sweet-waxy plants by primer phi061 in F2 populations obtained from L35×M01
From one to 21 indicate the PCR amplification products from F2 population genomic DNA obtained from L35 × M01. 1, 8, 11 and 12 indicate their corresponded plants with double recessive sweet-waxy genotype; M01: Sweet corn parental line, L35: Waxy corn parental line
表 1 鉴定wxwx基因型的3个SSR引物序列
Table 1 The sequences of three SSR primers for wxwxgenotype identification
引物名称
Primer name序列(5′→3′)
Primer sequencephi022 F-TGCGCACCAGCGACTGACC R-GCGGGCGACGCTTCCAAAC phi027 F-GCGTACGTACGACGAAGACAC R-CACAGCACGTTGCGGATTTCTCT phi061 F-AAACAAGAACGGCGGTGCTGATTC R-GACGTAAGCCTAGCTCTGCCAT 
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表 2 引物phi061在F2代群体中的χ2分析
Table 2 The chi-square analysis of primer phi061 in F2 populations
杂交组合
Hybrid combination植株总数量
Total plant number甜糯双隐性植株数量
No. of plants with wxwxsh2sh2非甜糯双隐性植株数量
No. of plants with Wx_sh2sh2χ2 P L33×M01 11 3 8 0.03 0.86 L35×M01 21 4 17 0.40 0.53 L38×M01 33 8 25 0.01 0.92
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表 3 11份甜糯双隐性自交系材料与甜、糯质亲本可溶性糖含量
Table 3 Soluble sugar contents in 11 double recessive sweet-waxy inbred lines and their parents
株系
Strain材料来源
Material sourcew(可溶性糖)1)/%
Soluble sugar content标准差/%
Standard deviation变异系数
Coefficient of variation变幅/%
RangeL1 L33×M01 13.44±1.63 2.82 0.21 11.03~16.54 L2 L33×M01 9.74±0.55 0.94 0.10 8.65~10.30 L3 L33×M01 12.04±1.02 1.76 0.15 10.00~13.08 L4 L35×M01 13.76±1.91 3.31 0.24 9.96~15.97 L5 L35×M01 10.73±1.01 1.74 0.16 8.95~12.43 L6 L35×M01 14.15±0.48 0.84 0.06 13.43~15.07 L7 L38×M01 13.88±1.45 2.51 0.18 12.27~16.77 L8 L38×M01 10.07±0.30 1.05 0.10 9.25~11.25 L9 L38×M01 12.42±0.29 0.50 0.04 11.89~12.87 L10 L38×M01 11.11±0.61 1.06 0.10 9.89~11.81 L11 L38×M01 13.59±0.69 1.19 0.09 11.25~14.49 M01 甜质亲本 9.72±0.39 0.67 0.07 9.25~10.49 L33 糯质亲本 3.76±0.35 0.60 0.16 3.08~4.19 L35 糯质亲本 5.13±0.85 1.48 0.29 3.97~6.78 L38 糯质亲本 1.64±0.51 0.51 0.31 1.15~2.18 1) 该列数据为平均值 ± 标准误
1) Data in this column are average value ± standard error
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表 4 11份甜糯双隐性自交系材料与甜、糯质亲本可溶性糖含量差异t检验1)
Table 4 The t test of differences in soluble sugar contents between 11 double recessive sweet-waxy inbred lines and their parents
株系
Strain甜质亲本 Sweet parent 糯质亲本 Waxy parent t P t P L1 3.14* 0.02 8.23** <0.01 L2 0.04 0.97 13.07** <0.01 L3 3.02* 0.02 7.37** <0.01 L4 2.93* 0.03 5.84** <0.01 L5 1.33 0.23 6.01** <0.01 L6 10.12** <0.01 31.23** <0.01 L7 3.91** <0.01 11.69** <0.01 L8 0.68 0.52 17.69** <0.01 L9 7.91** <0.01 36.88** <0.01 L10 2.72* 0.03 19.74** <0.01 L11 6.94** <0.01 22.57** <0.01 1) “*” 和 “**” 分别表示在 0.05 和 0.01 水平差异显著
1) “*” and “**” indicate significant differences at 0.05 and 0.01 levels, respectively
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表 5 11份甜糯双隐性自交系材料与甜、糯质亲本淀粉含量
Table 5 Starch contents in 11 double recessive sweet-waxy inbred lines and their parents
株系
Strain材料来源
Material sourcew(淀粉)1)/%
Starch content标准差/%
Standard deviation变异系数
Coefficient of variation变幅/%
RangeL1 L33×M01 21.97±1.07 1.86 0.08 19.92~23.51 L2 L33×M01 19.03±0.84 1.46 0.08 17.42~20.20 L3 L33×M01 21.13±1.18 2.04 0.10 18.89~22.88 L4 L35×M01 21.80±1.63 2.83 0.13 19.82~23.78 L5 L35×M01 17.90±0.78 1.35 0.08 16.77~23.04 L6 L35×M01 22.50±0.42 0.72 0.03 21.92~23.30 L7 L38×M01 19.23±1.05 1.81 0.09 17.33~20.88 L8 L38×M01 16.60±0.47 0.82 0.05 15.70~18.32 L9 L38×M01 20.63±0.75 1.31 0.06 18.60~22.11 L10 L38×M01 18.47±0.52 0.91 0.05 15.67~19.24 L11 L38×M01 17.80±0.79 1.37 0.08 16.30~19.01 M01 甜质亲本 23.50±0.99 1.71 0.07 21.92~25.31 L33 糯质亲本 50.20±4.10 7.11 0.14 42.92~57.08 L35 糯质亲本 46.33±1.86 3.22 0.07 42.88~49.03 L38 糯质亲本 45.50±3.13 5.43 0.12 41.59~51.72 1) 该列数据为平均值 ± 标准误
1) Data in this column are average value ± standard error
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表 6 11份甜糯双隐性自交系材料与甜、糯质亲本淀粉含量差异t检验1)
Table 6 The t test of differences in starch contents between 11 double recessive sweet-waxy inbred lines and their parents
株系
Strain甜质亲本 Sweet parent 糯质亲本 Waxy parent t P t P L1 –1.05 0.35 –6.65** <0.01 L2 –3.44* 0.03 –7.44** <0.01 L3 –1.54 0.20 –11.45** <0.01 L4 –0.83 0.45 –10.84** <0.01 L5 –4.46* 0.01 –14.11** <0.01 L6 –0.93 0.40 –7.27** <0.01 L7 –2.96* 0.04 –7.95** <0.01 L8 –3.96* 0.02 –9.42** <0.01 L9 –2.31 0.08 –7.71** <0.01 L10 –4.51* 0.01 –8.51** <0.01 L11 –4.50* 0.01 –8.56** <0.01 1) “*” 和 “**” 分别表示在 0.05 和 0.01 水平差异显著
1) “*” and “**” indicate significant differences at 0.05 and 0.01 levels, respectively
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表 7 11份甜糯双隐性自交系与甜、糯质亲本籽粒冠部和背胚侧果皮厚度1)
Table 7 The pericarp thickness on the crown and dorsal embryo side of the grain in 11 double recessive sweet-waxy inbred lines and their parents
株系
Strain材料来源
Material Source冠部 Crown 背胚侧 Dorsal embryo side 果皮厚度/μm
Pericarp thickness变幅/μm
Range果皮厚度/μm
Pericarp thickness变幅/μm
RangeL1 L33×M01 46.93±1.98 40.59~52.01 58.08±2.61 52.20~66.22 L2 L33×M01 35.29±0.81 33.09~37.53 43.07±1.51 39.32~46.86 L3 L33×M01 32.48±0.83 29.95~34.22 43.30±0.85 41.34~46.23 L4 L35×M01 43.02±1.19 39.49~46.77 52.56±0.63 50.26~53.94 L5 L35×M01 39.17±0.85 37.08~42.18 78.33±0.62 76.80~80.22 L6 L35×M01 39.04±0.31 38.06~40.03 61.04±0.92 58.29~63.37 L7 L38×M01 34.08±1.44 31.48~39.33 66.87±0.83 64.62~68.73 L8 L38×M01 46.75±1.60 41.97~51.20 64.23±0.66 62.53~66.17 L9 L38×M01 40.98±0.91 38.94~44.31 61.01±1.10 58.68~64.38 L10 L38×M01 43.17±2.46 38.66~49.35 60.23±2.49 52.98~65.81 L11 L38×M01 48.17±1.99 40.60~51.52 67.76±1.84 61.53~73.06 M01 甜质亲本 28.07±1.56 24.19~32.89 38.02±0.45 37.02~39.24 L33 糯质亲本 62.69±0.68 61.24~64.76 82.91±3.70 75.56~95.81 L35 糯质亲本 63.87±0.76 62.12~65.93 95.63±1.86 89.29~100.35 L38 糯质亲本 54.45±1.43 50.71~57.60 64.98±1.68 59.95~69.55 1) 表中数据为平均值 ± 标准误
1) Data in the table are average value ± standard error
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