Khatri Poonam, Joshee Nirmal
Agricultural Research Station, Fort Valley State University, Fort Valley, GA 31030, USA.
Plants (Basel). 2024 Jan 5;13(2):151. doi: 10.3390/plants13020151.
An efficient and reproducible in vitro method for indirect somatic embryogenesis was optimized by culturing leaf and leaf with petiole explants of L. Murashige and Skoog (MS) medium, supplemented with various concentrations of Picloram and 2,4-Dichlorophenoxyacetic acid (2,4-D), individually and in combinations, were tested. Picloram (1.0 µM) showed a better response compared to 2,4-D and results indicate it to be a better auxin for induction of somatic embryos for Goji berry. It was seen that the leaf explants were more responsive in callus and somatic embryo induction than the leaf with petiole explant when incubated in the dark for 5 weeks. Embryogenic callus, after being transferred to MS medium containing Benzyl amino purine (BAP) in 1.0 µM, 5.0 µM and 10.0 µM, began to differentiate in light after one week. MS medium with 1.0 µM Picloram + 10 µM BAP resulted as the most favorable treatment for somatic embryogenesis in L. Removal of plant growth regulators from MS medium and culturing induced calluses under 16 h photoperiod resulted in globular, heart, torpedo, cotyledons, and further development into plantlets. Well-developed plants have been obtained and are capable of acclimatizing in ex vitro conditions. In addition, the effects of desiccation treatments (0, 1, 3, 6, 9 h, and 12 h) on embryogenic callus for somatic embryo induction were found to be directly proportional to the length of desiccation treatment at room temperature. After 9 h and 12 h of desiccation treatments, 60% and 90% of plated calluses resulted in somatic embryos, respectively. In a callus mass, Acetocarmine and Evans blue double staining differentiated between embryogenic and non-embryogenic callus. These findings will help Goji berry improvement by elite clone production, ex situ conservation projects, scaling up plant production, and agronomy for the commercial production of this superfruit in the future.
通过在添加了不同浓度的毒莠定和 2,4 - 二氯苯氧乙酸（2,4 - D）的 Murashige 和 Skoog（MS）培养基上培养枸杞叶片和带叶柄叶片外植体，优化了一种高效且可重复的间接体细胞胚胎发生的体外方法，分别测试了单独使用和组合使用这两种植物生长调节剂的效果。与 2,4 - D 相比，毒莠定（1.0 µM）表现出更好的响应，结果表明它是诱导枸杞体细胞胚胎的更好生长素。可以看到，在黑暗中培养 5 周时，叶片外植体在愈伤组织和体细胞胚胎诱导方面比带叶柄叶片外植体更敏感。将胚性愈伤组织转移到含有 1.0 µM、5.0 µM 和 10.0 µM 苄氨基嘌呤（BAP）的 MS 培养基中，一周后在光照下开始分化。含有 1.0 µM 毒莠定 + 10 µM BAP 的 MS 培养基是枸杞体细胞胚胎发生最有利的处理方法。从 MS 培养基中去除植物生长调节剂并在 16 小时光周期下培养诱导的愈伤组织，会形成球形、心形、鱼雷形、子叶形，并进一步发育成小植株。已经获得了发育良好的植株，并且它们能够在体外条件下适应环境。此外，发现在室温下对胚性愈伤组织进行干燥处理（0、1、3、6、9 小时和 12 小时）对体细胞胚胎诱导的影响与干燥处理时间长度成正比。经过 9 小时和 12 小时的干燥处理后，分别有 60%和 90%接种的愈伤组织产生了体细胞胚胎。在愈伤组织团中，醋酸洋红和伊文思蓝双重染色可区分胚性愈伤组织和非胚性愈伤组织。这些发现将有助于通过优良克隆生产、迁地保护项目、扩大植物生产以及未来这种超级水果商业化生产的农艺学来改良枸杞。
