Aroma is a key grain quality trait that directly influences the market price of rice globally. Loss of function of betaine aldehyde dehydrogenase 2 (OsBADH2) affects the biosynthesis of 2-acetyl-1-pyrroline (2-AP), which is responsible for aroma in fragrant rice. The current study was aimed at creating new alleles of BADH2 using CRISPR/Cas9 gene editing technology under the genetic background of the japonica Ningjing 1 (NJ1) and indica Huang Huazhan (HHZ) varieties. Sensory evaluation and analysis using headspace solid-phase microextraction gas chromatography–mass spectrometry (HS-SPME-GC-MS) showed that the grains of the four homozygous T₁ lines with new alleles of BADH2 (nj1-cr ^BADH2-1, nj1-cr ^BADH2-2, hhz-cr ^BADH2-1 and hhz-cr ^BADH2-2) produced moderate fragrance and had significantly increased 2-AP content compared with wild-types. Moreover, there were no significant differences in the amylose content and gelatinization temperature among the four lines with new alleles of BADH2 to the wild-types. Thereafter, we crossed the HHZ background new alleles of BADH2 with CMS line Taonong 1A (TN1A) to produce a three-line hybrid variety B-Tao-You-Xiangzhan (BTYXZ) with increased grain aroma. The 2-AP content in grains of the improved BTYXZ-1 and BTYXZ-2 reached at 26.16 and 18.74 μg/kg, and the gel consistency of BTYXZ-1 and BTYXZ-2 increased significantly by 9.1% and 6.5%, respectively, compared with the wild-type Tao-You-Xiangzhan (TYXZ). However, the γ-aminobutyric acid (GABA) content in the improved three-line hybrid rice BTYXZ-1 (5.6 mg/100 g) and BTYXZ-2 (10.7 mg/100 g) was significantly lower than that of the TYXZ. These results demonstrated that CRISPR/Cas9 gene editing technology could be successfully utilized in improving aroma in non-fragrant japonica and indica varieties. In addition, the newly developed BADH2 alleles provided important genetic resources for grain aroma improvement in three-line hybrid rice.

中文翻译：

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BADH2新等位基因生产芳香三系杂交水稻

香气是直接影响全球大米市场价格的关键谷物品质性状。甜菜碱醛脱氢酶 2 ( OsBADH2 )的功能丧失会影响 2-乙酰基-1-吡咯啉 (2-AP) 的生物合成，而 2-AP 负责香米的香气。本研究旨在在粳稻宁靖1号（NJ1）和籼稻黄华占（HHZ）品种的遗传背景下，利用CRISPR/Cas9基因编辑技术创造新的BADH2等位基因。使用顶空固相微萃取气相色谱-质谱 (HS-SPME-GC-MS) 进行感官评价和分析表明，具有 BADH2 新等位基因 (nj1-cr BADH2 -1) 的四个纯合T 1 ^系的_颗粒, nj1-cr  ^BADH2 -2 , hhz-cr  ^BADH2 -1和hhz-cr  ^BADH2 -2 ) 与野生型相比，香气适中，2-AP含量显着增加。此外，与野生型有新等位基因BADH2的4个品系的直链淀粉含量和糊化温度无显着差异。此后，我们越过了BADH2的 HHZ 背景新等位基因与 CMS 系桃农 1A (TN1A) 生产具有增强谷物香气的三系杂交品种 B-桃油-香展 (BTYXZ)。改良后的 BTYXZ-1 和 BTYXZ-2 籽粒中 2-AP 含量分别达到 26.16 和 18.74 μg/kg，BTYXZ-1 和 BTYXZ-2 的凝胶稠度分别显着提高了 9.1% 和 6.5%。与野生型桃有象展（TYXZ）。但改良三系杂交稻BTYXZ-1（5.6 mg/100 g）和BTYXZ-2（10.7 mg/100 g）的γ-氨基丁酸（GABA）含量明显低于TYXZ。这些结果表明，CRISPR/Cas9基因编辑技术可以成功地用于改善粳稻和籼稻品种的香气。此外，新开发的BADH2等位基因为三系杂交水稻籽粒香气改良提供了重要的遗传资源。