Embryo Rescue of Interspecific Brassica campestris x B. albograbra Hybrids
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摘要:目的 获得适合菜薹和芥蓝种间杂种的胚挽救实验方案。方法 以菜薹（菜心）作为母本，芥蓝作为父本，进行种间杂交，对种间杂种进行人工胚挽救培养，设置8种不同成分的胚挽救培养基，6个不同取材时间，4种增殖培养基，9种不同成分生根培养基。结果 ERM4（MS+6-BA 0.5 mg·L−1+NAA 0.5 mg·L−1+KT 1.0 mg·L−1+AC 1.0 g·L−1）出胚率达26.32%，为最佳胚挽救培养基；授粉后第12天取材，出胚率达26.32%，为最适宜取材时间；DM1（MS+6-BA 0.9 mg·L−1+NAA 0.1 mg·L−1）培养基增殖系数最大，为5.37，为最佳增殖培养基；RM4-2（1/2MS+IBA0.5 mg·L−1）培养基生根率为37.21%，平均生根时间为8.33 d，生根最快，为最佳生根培养基。结论 菜薹与芥蓝种间杂交，于授粉后12 d取材，胚珠接种在ERM4培养基上，出胚后接种在DM1培养基上增殖，在RM4-2培养基上生根，能够显著提高种间杂种的获得率。Abstract:Objective Methods suitable for embryo rescue in the interspecific hybridization between Brassica campestris and B. albograbra were investigated.Method The interspecific crossing was carried out using B. campestris L. as female parent and B. albograbra L. H. Bailey as male parent. An embryo rescue culture for the hybridization was conducted with 8 different media at 6 sampling times followed subsequently by 4 proliferation media and 9 rooting media.Result The embryo genesis rate at 26.32% of ERM4 medium consisting of MS, 0.5mg 6-BA·L−1, 0.5mg NAA·L−1, 1.0mg KT·L−1, and 1.0g AC·L−1 was the highest among all. The samples collected 12d after pollination had the best rate of 26.32%, DM1 (i.e., MS+ 0.9mg 6-BA·L−1+0.1mg NAA·L−1) the greatest proliferation coefficient of 5.37, and RM4-2 (i.e., 1/2MS+ 0.5mg IBA·L−1) the top rate of 37.21% and average 8.33d on rooting.Conclusion The optimum conditions for the embryo rescue in the interspecific hybridization between B. campestris L. and B. albograbra L. H. Bailey included sampling at 12d after pollination and following by culture on ERM4 medium for embryo induction, on DM1 for ovule proliferation, and on RM4-2 for seedling rooting.
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图 1 菜薹×芥蓝种间杂种胚挽救情况
注：A，胚珠萌发；B，胚挽救幼苗；C，成活胚增殖分化
Figure 1. Embryo rescue of interspecific B. campestris × B. albograbra hybrids
Note: A: Ovule germination; B: embryo rescued seedling; C: proliferation and differentiation of survival embryos.
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图 2 菜薹×芥蓝种间杂种生根过程不同生长素生长果
注：A：NAA生根效果；B：IBA生根效果；C：IAA生根效果；D：未加激素MS培养基生根效果
Figure 2. Effects of auxins on growth during rooting of interspecific B. campestris × B. albograbra hybrids
Note: A: Rooting affected by NAA; B: rooting affected by IBA; C: rooting affected by IAA; D: seedling rooting on MS without hormone addition.
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图 3 菜薹×芥蓝种间杂种生根过程盐害现象
Figure 3. Damage induced by salt at rooting stage of interspecific B. campestris × B. albograbra hybrids
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图 4 菜薹×芥蓝种间杂种生根过程
Figure 4. Rooting of interspecific B. campestris × B. albograbra hybrids
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图 5 菜薹×芥蓝种间杂种植株形态
注：A，菜薹；B、C，种间杂种；D，芥蓝
Figure 5. Morphology of interspecific B. campestris × B. albograbra hybrid plant
Note：A: B. campestris; B and C: interspecific hybrid; D: B. albograbra.
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图 6 菜薹×芥蓝种间杂种花器官形态
注：A：F菜薹；B：E种间杂种；C：D芥蓝
Figure 6. Morphology of floral organ of interspecific B. campestris × B. albograbra hybrid
Note: A and F: B. campestris; B and E: interspecific hybrid; C and D: B. albograbra.
表 1 菜薹×芥蓝种间杂种胚挽救培养基成分
Table 1 Components in embryo rescue medium for interspecific B. campestris × B. albograbra hybrids
编号
No.基本培养基
Minimal medium6-BA质量浓度
6-BA mass concentration/
（mg·L−1）NAA质量浓度
NAA mass concentration/
（mg·L−1）KT质量浓度
KT mass concentration/
（mg·L−1）AC含量
AC content/
（g·L−1）ERM1 MS 0.00 0.00 0.00 1.00 ERM2 MS 0.30 0.00 0.00 1.00 ERM3 MS 0.50 0.00 0.00 1.00 ERM4 MS 0.50 0.50 1.00 1.00 ERM5 MS 1.50 0.50 1.00 1.00 ERM6 MS 3.00 0.50 1.00 1.00 ERM7 MS 4.50 0.50 1.00 1.00 ERM8 MS 5.00 0.50 1.00 1.00 
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表 2 菜薹×芥蓝种间杂种增殖培养基成分
Table 2 Components in propagation medium for interspecific B. campestris × B. albograbra hybrids
编号
No.基本培养基
Minimal
medium6-BA质量浓度
6-BA mass
concentration/
（mg·L−1）NAA质量浓度
NAA mass
concentration/
（mg·L−1）DM1 MS 0.90 0.10 DM2 1/2MS 0.90 0.10 DM3 MS 0.50 0.01 DM4 1/2MS 0.50 0.01
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表 3 菜薹×芥蓝种间杂种生根培养基成分
Table 3 Components in rooting medium for interspecific B. campestris × B. albograbra hybrids
编号
No.基本培养基
Minimal
mediumNAA质量
浓度
NAA mass concentration/
（mg·L−1）IBA质量
浓度
IBA mass concentration/
（mg·L−1）IAA质量
浓度
IAA mass concentration/
（mg·L−1）RM1-1 MS 0.20 0.00 0.00 RM1-2 0.50 0.00 0.00 RM2-1 0.00 0.20 0.00 RM2-2 0.00 0.50 0.00 RM3-1 0.00 0.00 0.20 RM3-2 0.00 0.00 0.50 RM4-1 1/2MS 0.20 0.00 0.00 RM4-2 0.00 0.50 0.00 RM4-3 0.00 0.00 0.50
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表 4 菜薹×芥蓝杂种胚珠在不同培养基上出胚率
Table 4 Embryogenesis rate of B. campestris × B. albograbra hybrid ovules on different media
编号
No.胚珠培养数
Number of
ovules成胚数
Number of
embryogenesis出胚率
Embryogenesis
rate/%ERM1 105 0 0.00±0.00 f ERM2 75 1 1.33±0.02 ef ERM3 78 2 2.56±1.25 e ERM4 95 25 26.32±0.78 a ERM5 88 16 18.18±0.93 b ERM6 85 13 15.29±1.00 c ERM7 101 10 9.90±0.89 d ERM8 103 8 7.77±0.90 d 注：不同小写字母表示差异达0.05显著水平。下同。
Note: Different lowercase letters indicate significant difference at P<0.05. Same for the following.
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表 5 菜薹×芥蓝不同取材时间杂种胚珠出胚率
Table 5 Embryogenesis rate of B. campestris × B. albograbra hybrid ovules with differed sampling time
授粉后天数
Days after
pollination/d胚珠培养数
Number of
ovules出胚数
Number of
embryogenesis出胚率
Embryogenesis
rate/%10 90 4 4.44±1.93 b 11 88 9 10.23±1.97 b 12 95 25 26.32±2.77 a 13 382 40 10.47±0.92 b 14 150 10 6.67±3.34 b 15 60 2 3.33±2.89 c
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表 6 菜薹×芥蓝种间杂种成活胚珠在不同培养基上增殖分化情况
Table 6 Proliferation and differentiation of viable ovules in B. campestris × B. albograbra hybrids on different media
编号
No.原有芽数
Number of
original bud出芽数
Number of
budding增殖系数
Multiplication
coefficientDM1 79 424 5.37±0.05 a DM2 76 306 4.02±0.38 b DM3 80 332 4.15±0.25 b DM4 78 226 2.90±0.15 c
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表 7 菜薹×芥蓝种间杂种不同培养基生根情况
Table 7 Rooting of interspecific B. campestris × B. albograbra hybrids on different media
编号
No.培养幼苗数
Number of
cultured seedlings生根幼苗数
Number of
rooting seedlings生根率
Rate of
rooting/%平均生根时间
Mean rooting
time/d移栽后成活幼苗
Number of survived
seedlings after
transplanting移栽成活率
Rate of
transplanting
survival/%RM1-1 27 11 40.74±2.20 ab 11.00±1.00 b 11 100 RM1-2 28 4 14.29±3.07 e 11.33±0.58 b 4 100 RM2-1 24 6 25.00±3.13 cd 8.67±0.58 b 6 100 RM2-2 25 9 36.00±2.57 bc 8.67±0.58 b 9 100 RM3-1 27 5 18.52±3.03 d 14.33±1.53 a 5 100 RM3-2 27 7 25.93±2.75 c 15.00±1.00 a 7 100 RM4-1 45 20 44.44±1.93 a 11.33±0.58 b 20 100 RM4-2 43 16 37.21±1.47 cd 8.33±0.58 c 16 100 RM4-3 47 13 27.66±1.54 c 14.67±1.53 a 13 100
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