NMT是基因功能的活体检测技术，已被103位诺贝尔奖得主所在单位，及北大、清华、中科院使用。
期刊：International Journal of Molecular Sciences
主题：OsCIPK2过表达促进低氮条件下水稻NO3-吸收
标题：Exploring the Potential of Overexpressed OsCIPK2 Rice as a Nitrogen Utilization E cient Crop and Analysis of Its Associated Rhizo-Compartmental Microbial Communities
影响因子：4.183
检测指标：Ca2+、NH4+、NO3-流速
检测样品：水稻根尖
Ca2+、NH4+、NO3-流实验处理方法：
幼苗长至三叶期，分别在正常氮（1.44 mM NH4NO3）和低氮（0.24 mM NH4NO3）处理
Ca2+、NH4+、NO3-流实验测试液成份：
Ca2+: 0.1 mmol/L KCl, 0.1 mmol/L CaCl2, 0.1 mmol/L MgCl2, 0.5 mol/L NaCl, 0.3 mmol/L MES, 0.2 mmol/L Na2SO4, pH 6.0;
NO3-: 0.1 mmol/L NH4NO3, 0.1 mmol/L NH4NO3, 0.1 mmol/L KCl, 0.1 mmol/L KCl, 0.1 mmol/L KCl2, 0.3 mmol/CaCl2, 0.3 mmol/L MgCl2, 0.3 mmol；
NH4+: 0.1 mmol/L NH4NO3, 0.1 mmol/L CaCl2, 0.3 mmol/L MES, pH 6.0
作者：福建农林大学林文雄
英文摘要
Nitrogen (N) is one of the indispensable factors in rice growth and development. China holds a premier position in the production of rice and at the same time also faces higher N fertilizer costs along with serious damage to the environment. A better solution is much needed to address these issues, without disrupting the production of rice as an important cereal, while minimizing all the deleterious effects on the environment.
Two isogenic lines Kitaake (WT) and its genetically modified line CIPK2 (RC), overexpressing the gene for Calcineurin B-like interacting protein kinase 2 (OsCIPK2) with better nitrogen use efficiency (NUE), were compared for their growth and development under low versus normal levels of N. NUE is a complex trait mainly related to a plant’s efficiency in extraction, assimilation, and recycling of N from soil. The microbial population was analyzed using high-throughput Illumina Miseq 16S rRNA sequencing and found that RC with CIPK2, specifically expressed in rice root, not only performed better without nitrogen fertilizer (LN) but also increased the diversity of bacterial communities in rice rhizosphere compartments (rhizosphere, rhizoplane, and endosphere).
The relative abundance of beneficial bacteria phyla increased, which are known to promote the circulation and transformation of N in rhizosphere soil. To further explore the potential of RC regarding better performance under LN, the ion fluxes in root apical were detected by non-invasive micro-test technique (NMT). We found that RC can absorb more Ca2+ and NO3− under LN as compared to WT. Finally, compared to WT, RC plants exhibited better growth of root and shoot, and increased yield and N uptake under LN, whereas there was no significant difference in the growth of two rice lines under normal nitrogen (NN) treatment.
We are able to get preliminary results, dealing with the OsCIPK2 overexpressed rice line, by studying the rice molecular, physiological, and chemical parameters related to NUE. The results laid the foundation for further research on N absorption and utilization in rice from the soil and the interaction with microbial communities.
中文摘要（谷歌机翻）
氮（N）是水稻生长发育不可或缺的因素之一。中国在大米生产中处于**地位，同时也面临着更高的氮肥成本以及对环境的严重破坏。迫切需要一种更好的解决方案来解决这些问题，同时又不中断作为重要谷物的大米的生产，同时将对环境的所有有害影响降至最低。
比较了两个同基因系Kitaake（WT）及其基因改造品系CIPK2（RC），它们以较低的氮素利用效率（NUE）过量表达了钙调磷酸酶B样相互作用蛋白激酶2（OsCIPK2）的基因，并在低温下对其生长发育进行了比较。与正常水平的氮相比。NUE是一个复杂的性状，主要与植物从土壤中提取，吸收和循环利用N的效率有关。使用高通量Illumina Miseq 16S rRNA测序分析了微生物种群，发现带有CIPK2的RC在水稻根部特异表达，不仅在不使用氮肥（LN）的情况下表现更好，而且增加了水稻根际区室细菌群落的多样性（根际，根际和内球）。
有益菌门的相对丰度增加，已知它可以促进根际土壤中氮的循环和转化。为了进一步探索RC在LN下获得更好性能的潜力，通过无创微测试技术（NMT）检测了根尖中的离子通量。我们发现，与野生型相比，RC在LN下可以吸收更多的Ca2+和NO3-。最后，与WT相比，RC植物在LN下表现出更好的根和茎生长，并增加了产量和氮素吸收，而在正常氮（NN）处理下，两个水稻品系的生长没有显着差异。
通过研究与NUE相关的水稻分子，生理和化学参数，我们能够获得初步结果，处理过表达OsCIPK2的水稻品系。研究结果为进一步研究水稻从土壤中吸收和利用氮以及与微生物群落的相互作用奠定了基础。
结果表明：LN（low nitrogen）处理时，RC（过表达型）根系的Ca2+吸收显著增加，而NN（normal nitrogen）处理时无显著差异；LN处理时，RC根系的NO3-吸收增加，而NN条件下RC和WT（野生型）根系的NO3-表现为外排，且二者并无显著差异；至于NH4+，RC和WT均表现为外排，且无显著差异。这说明，LN条件下，根系CIPK2基因过表达促进了根系吸收NO3-，与RC根际的硝化作用增加是符合的。