Improving Blast Resistance Of Rice CMS Line SanxiangA Through Marker-assisted Selection Of The Pi9 Gene

Rice（Oryza sativa）is one of the world’s most important food crops.Rice blast is one of the most serious diseases affecting rice production during the whole cycle of rice growth and development.The most effective way to prevent and control rice blast is to breed varieties with high rice blast resistance.In this study,indica rice line 75-1-127 was used as the donor parent of Pi9 gene for broad spectrum durable resistance to rice blast,and the indica CMS line SanxiangA and its maintainer SanxiangB were used as the recipient parents.To improve blast resistance of SanxiangA and SanxiangB,the codominant In De L marker CoInDF₁R₁was used in marker-assisted selection backcross breeding.The main results were as follows:1.Blast resistance of parental rice lines was investigated by means of greenhouse inoculation and field nursery assays.Resistance responses to 28 tested M.oryzae isolates showed obvious difference between 75-1-127 and two receptor parents,presenting 85.71%and 28.57%resistance frequency respectively,and overlapping resistance-spectrum was observed between the donor and receptors.Meanwhile,75-1-127 showed high-level seedling blast resistance（0 grade）but highly susceptible（4～5 grades）for two receptor parents in three years’consecutive natural nursery blast resistance identification.These results indicated that Pi9 gene possesses broad-spectrum and high-level blast resistance with huge potential in breeding practice,while the receptor parents SanxiangA and SanxiangB showed narrow-spectrum and low-level blast resistance and need to be improved.2.Ten functional molecular markers were developed based on DNA sequence information of the Pi9 gene,and one of them,CoInDF₁R_1,showed obvious and stable polymorphism between the donor/receptor parents.The CoInDF₁R₁marker primers amplified a 320bp band from the genome of 75-1-127 while the amplification product from genome of SanxiangA or Sanxianng B was about 410bp.Furthermore,this codominant marker can be analyzed simply by agarose gel electrophoresis.3.CoInDF₁R₁was then employed for marker-assisted selection（MAS）backcross breeding practice to improve blast resistance of SanxiangA and its maintainer SanxiangB in this study.Twelve improved maintainer lines SanxiangB-Pi9 and nine improved CMS lines SanxiangA-Pi9 were developed,which carrying the Pi9 homozygous alleles and showing high-level seedling blast resistance.4.The improved CMS lines SanxiangA-Pi9-3,SanxiangA-Pi9-6,SanxiangA-Pi9-9 and their maintainers SanxiangB-Pi9-3,SanxiangB-Pi9-6,and SanxiangB-Pi9-9 were highly resistant to M.oryzae isolate E2007046A2 in greenhouse spray inoculation assay as their donor parent 75-1-127.Meanwhile,above improved CMS lines and their maintainers showed identical seedling blast resistance as 75-1-127 either in in-vitro inoculation by M.oryzae isolates E2007046A2,110-2 and X2007A-1,or in natural nusery investigation.These results showed that seedling blast resistance of SanxiangA and its maintainer SanxiangB was completely improved through MAS breeding in this study.5.Sterility and cross-pollination traits of three improved CMS lines,and main agronomic and yield traits of the three CMS lines and the corresponding maintainers were investigated by field cultivation assays.Finally,an elite CMS line SanxiangA-Pi9-3 and its maintainer SanxiangB-Pi9-3 were bred.SanxiangA-Pi9-3 showed high-level seedling blast resistance,complete male sterility,high stigma exsertion rate（61.7%）and better yield trait.These work paved a way for further breeding novel hybrids with strong heterosis through three-lines method.