Method for breeding yellow-seeded brassica napus glyphosate-resistant recessive genic male sterility temporary maintainer line
A glyphosate-resistant Brassica napus technology, which is applied in the field of crop breeding, can solve the problems of heavy test cross workload, heavy fertility identification of offspring, difficult field implementation, and large segregation groups.
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Embodiment 1
[0049] The culture medium that the embodiment of the present invention uses is as follows:
[0050] (1) Microspore isolation medium (B5-13)
[0051] B5 macroelements + B5 trace elements + B5 organic additives + FeNaEDTA 13.2mg L -1 + sucrose 130g·L -1 (pH=6.0)
[0052] (2) embryoid body induction medium NLN-13, wherein, the composition of NLN-13 medium is as follows:
[0053] Macroelements: KNO 3 125mg·L -1 , MgSO 4 ·7H 2 O 125mg·L -1 , Ca 4 NO 3 .4H 2 O 500mg·L -1 、KH 2 PO 4 125mg·L -1 ;
[0054] Iron salt: FeNaEDTA 26.4mg·L -1 ;
[0055] Trace elements: MnSO 4 4H 2 O 22.3mg·L -1 、H 3 BO 3 6.2mg·L -1 , ZnSO 4 ·7H 2 O 8.6mg·L -1 , CuSO4 5H 2 O 0.025mg·L -1 、Na 2 MoO 4 2H 2 O 0.25mg·L -1 、CoCl 2 ·6H 2 O 0.025mg·L -1 ;
[0056] Organic additives: inositol 100mg·L -1 , Niacin 5mg·L -1 , pyridoxine hydrochloride 0.5mg·L -1 , Thiamine Hydrochloride 0.5mg·L -1 , Glycine 2mg·L -1 , folic acid 0.5mg·L -1 , Biotin 0.05mg·L -1 , Glutathione ...
Embodiment 2
[0064] In March 2018, at the flowering stage of rapeseed, WAL1A was used as the female parent and the male parent SWUWY-35 was crossed to obtain F1 generation seeds.
[0065] In March 2018 rape flowering period, for F 1 Microspore culture was carried out, and a total of 200 embryoid bodies were obtained, and 160 individual plants were obtained through induction culture. They were planted in the field in October 2018, and 88 doubled plants were identified in March 2019, and then identified by glyphosate and molecular markers 60 glyphosate-resistant plants were obtained. The specific process is as follows:
[0066] (1) From a robust plant F 1 Above, pick the flower buds of the main inflorescence or the top part of the primary branch inflorescence 3.5-4mm, soak in 75% alcohol for 0.5-1min, rinse with sterile water for 3 times, and then use 0.1% HgCl 2 Soak for 12 minutes, rinse with sterile water for 3 times, about 5 minutes each time;
[0067] (2) Transfer the sterilized flo...
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