Tang Weijie, Chen Haiyuan, Zhang Suobing, Tang Jun, Lin Jing, Fang Xianwen, Chen Gaoming, Zhang Yunhui
Provincial Key Laboratory of Agrobiology, Institute of Germplasm Resources and Biotechnology, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China.
Zhongshan Biological Breeding Laboratory, Nanjing 210014, China.
Plants (Basel). 2024 Mar 6;13(5):745. doi: 10.3390/plants13050745.
is the key gene that controls amylose content (AC), and various alleles have been found in rice populations. is the major allele in and produces moderate AC (15~18%). It was recently found that editing the promoter of could produce a series of alleles that have different Wx activities. Although some studies have edited the promoter, few studies have focused on the natural variations in . Here, we used the Rice3K database to investigate variations in the promoter and found that the allele Wx1764178 (A/G) has a higher LD (linkage disequilibrium) with the two key SNPs (1765751, T/G; 1768006, A/C), which could produce different alleles and influence AC, as reported previously. Further study showed that the Wx1764178 allele (A/G) is functional and influences the expression of positively. Editing the A allele using CRISPR‒Cas9 produced 36 and 3 bp deletions and caused a decrease in the expression of . The apparent amylose content (AAC) in the edited lines was decreased by 7.09% and 11.50% compared with that of the wild type, which was the variety Nipponbare with and the A allele at 1764178, while a complementary line with the G allele showed a lower AAC than the A allele with no effect on other agronomic traits. The AAC of the edited lines showed a higher increase than that of the wild type (Nipponbare, ) in low-nitrogen conditions relative to high-nitrogen conditions. We also developed a dCAPS marker to identify the allele and found that the G allele has widely been used (82.95%) in -bred varieties from Jiangsu Province, China. Overall, we found a functional allele (Wx1764178, A/G) in the promoter that could affect AAC in cultivars and be developed as markers for quality improvement in rice breeding programs.
是控制直链淀粉含量（AC）的关键基因，在水稻群体中已发现多种等位基因。是中的主要等位基因，产生中等AC（15%~18%）。最近发现，编辑的启动子可产生一系列具有不同Wx活性的等位基因。虽然一些研究对启动子进行了编辑，但很少有研究关注的自然变异。在此，我们利用Rice3K数据库研究启动子的变异，发现等位基因Wx1764178（A/G）与两个关键单核苷酸多态性（1765751，T/G；1768006，A/C）具有较高的连锁不平衡（LD），如先前报道，这两个关键单核苷酸多态性可产生不同的等位基因并影响AC。进一步研究表明，Wx1764178等位基因（A/G）具有功能且正向影响的表达。使用CRISPR-Cas9编辑A等位基因产生了36和3bp的缺失，并导致表达下降。与野生型相比，编辑品系中的表观直链淀粉含量（AAC）分别降低了7.09%和11.50%，野生型为具有且在1764178处为A等位基因的日本晴品种，而具有G等位基因的互补品系显示出比A等位基因更低的AAC，且对其他农艺性状无影响。相对于高氮条件，在低氮条件下，编辑品系的AAC比野生型（日本晴，）有更高的增加。我们还开发了一个dCAPS标记来鉴定该等位基因，发现G等位基因在中国江苏省的自交品种中广泛使用（82.95%）。总体而言，我们在启动子中发现了一个功能性等位基因（Wx1764178，A/G），其可影响水稻品种的AAC，并可开发为水稻育种计划中品质改良的标记。
