Method for breeding hybrid seeds of rape of heterogenesis cytoplasm male sterility line
A male sterile line and male sterile technology, applied in the field of plant bioengineering, can solve problems such as unstable fertility, decreased purity of hybrid seed production, and PolCMS is easily affected by temperature, achieving high induction success rate and hybrid selection Easy, stable and fast results
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Embodiment 1
[0076] Implementation Example 1: Intergeneric hybridization creates male sterility in Brassica napus containing wild mustard cytoplasm
[0077] 1. Use plant material to carry out protoplast fusion or sexual hybridization (see implementation example 2) to obtain a somatic cell hybrid containing wild mustard cytoplasm. The plant material used is Brassica napus double-low variety Zhongshuang 4 or Xinjiang wild mustard resource wild oil 18 number, including the following steps:
[0078] A., Isolation of mesophyll protoplasts: after surface disinfection (put 70% ethanol for 1 minute, 1.5% sodium hypochlorite for 10 minutes, rinse 3 times with sterile water), the seeds of Brassica napus and wild mustard are inoculated in MS On the medium, set 25oC, 16 hours 60-80μmol m -2 the s -1 Grow under light conditions. Fully expanded leaves were used for the isolation of protoplasts. During enzymatic hydrolysis, 3.5ml of SCM solution (0.5M sorbitol, 10mM CaCl2, 5mM MES, pH5.8) was added t...
Embodiment 2
[0120] Example 2: Obtaining Somatic Cell Hybrids Containing Mustard Cytoplasm by Sexual Crossing
[0121] 1) Ovary culture to produce intergenus hybrids containing wild mustard cytoplasm: plant wild mustard and Brassica napus in the field, take wild mustard as female parent and Brassica napus as male parent in flowering stage, carry out artificial emasculation and pollination, after pollination 7-10 days, harvest Back to the siliques that have begun to develop, after surface disinfection (putting 70% ethanol for 1 minute, 1.5% sodium hypochlorite for 10 minutes, and rinsing with sterile water for 3 times), put the MS medium containing 0.8% hydrolyzed casein 60-80μmol m at 25°C for 16 hours -2 the s -1 Cultivate under light conditions. After 35-40 days, take out the immature embryos in the siliques and germinate them on MS medium. After germination, 60-80 μmol m -2 the s -1 Let it grow true leaves under light conditions.
[0122] 2) Chromosome doubling of intergenus hybrids: ...
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