{"title":"桔梗原生质体的分离和培养:影响原生质体产量、细胞分裂和微球茎形成的因素","authors":"Suk-Hyun Kwon, Hosakatte Niranjana Murthy, Jong-Eun Han, Hon-Sol Lee, So-Young Park","doi":"10.1007/s13580-023-00585-0","DOIUrl":null,"url":null,"abstract":"<p><i>Plyatocodon grandiflorus</i> is an important medicinal herb, its rhizomes are utilized as vegetables and as medicinal food. In this investigation, we were able to successfully isolate protoplasts from <i>P. grandiflorus</i> leaf mesophyll tissues in order to promote the development of calli. It has been determined what are the ideal conditions for protoplast isolation, involving the combination of enzymes, the osmoticum, and the length of enzyme digestion. For the most effective results, cell protoplast washing (CPW) medium with 1.0% Viscozyme® L + 3% Celluclast® 1.5 L + 0.5% Pectinex® XXL + 0.4 M mannitol and 8 h of incubation was used to isolate protoplasts from leaf mesophyll tissues. The thin alginate layer (TAL) approach was utilized to embed the protoplasts. Subsequently, TAL segments were cultured in Murashige and Skoog (MS) media supplemented with 3% sucrose, 2,4-dichlorophenoxyacetic acid (2,4-D, 0, 0.56, 1.12 µM), and phytosulfokine (PSK, 0, 0.05, 0.1 µM). The optimal concentration of 2,4-D and PSK for initiation of protoplast division, multicell development, and micro-callus formation was predicted using response surface methodology (RSM) with central composite design (CCD). Based on the CCD responses, it was determined that 1.12 µM 2,4-D and 0.065 µM PSK were the best concentrations to induce protoplast division and the formation of micro-calluses. The acquired results are valuable for <i>P. grandiflorus</i> protoplast isolation and culture; however, coordinated efforts are required for plant regeneration using callus produced from protoplasts.</p>","PeriodicalId":13123,"journal":{"name":"Horticulture Environment and Biotechnology","volume":null,"pages":null},"PeriodicalIF":2.4000,"publicationDate":"2024-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Isolation, culture of Platycodon grandiflorus protoplasts: factors affecting protoplast yield, cell division, and micro-callus formation\",\"authors\":\"Suk-Hyun Kwon, Hosakatte Niranjana Murthy, Jong-Eun Han, Hon-Sol Lee, So-Young Park\",\"doi\":\"10.1007/s13580-023-00585-0\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><i>Plyatocodon grandiflorus</i> is an important medicinal herb, its rhizomes are utilized as vegetables and as medicinal food. In this investigation, we were able to successfully isolate protoplasts from <i>P. grandiflorus</i> leaf mesophyll tissues in order to promote the development of calli. It has been determined what are the ideal conditions for protoplast isolation, involving the combination of enzymes, the osmoticum, and the length of enzyme digestion. For the most effective results, cell protoplast washing (CPW) medium with 1.0% Viscozyme® L + 3% Celluclast® 1.5 L + 0.5% Pectinex® XXL + 0.4 M mannitol and 8 h of incubation was used to isolate protoplasts from leaf mesophyll tissues. The thin alginate layer (TAL) approach was utilized to embed the protoplasts. Subsequently, TAL segments were cultured in Murashige and Skoog (MS) media supplemented with 3% sucrose, 2,4-dichlorophenoxyacetic acid (2,4-D, 0, 0.56, 1.12 µM), and phytosulfokine (PSK, 0, 0.05, 0.1 µM). The optimal concentration of 2,4-D and PSK for initiation of protoplast division, multicell development, and micro-callus formation was predicted using response surface methodology (RSM) with central composite design (CCD). Based on the CCD responses, it was determined that 1.12 µM 2,4-D and 0.065 µM PSK were the best concentrations to induce protoplast division and the formation of micro-calluses. The acquired results are valuable for <i>P. grandiflorus</i> protoplast isolation and culture; however, coordinated efforts are required for plant regeneration using callus produced from protoplasts.</p>\",\"PeriodicalId\":13123,\"journal\":{\"name\":\"Horticulture Environment and Biotechnology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.4000,\"publicationDate\":\"2024-01-15\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Horticulture Environment and Biotechnology\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://doi.org/10.1007/s13580-023-00585-0\",\"RegionNum\":3,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"Agricultural and Biological Sciences\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Horticulture Environment and Biotechnology","FirstCategoryId":"97","ListUrlMain":"https://doi.org/10.1007/s13580-023-00585-0","RegionNum":3,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"Agricultural and Biological Sciences","Score":null,"Total":0}
引用次数: 0
摘要
桔梗是一种重要的药材,其根茎可用作蔬菜和药用食品。在这项研究中,我们成功地从大叶杓兰叶叶肉组织中分离出原生质体,以促进胼胝体的发育。我们已经确定了原生质体分离的理想条件,包括酶的组合、渗透压和酶消化时间的长短。为了获得最有效的结果,使用了细胞原生质体洗涤(CPW)培养基,即 1.0% Viscozyme® L + 3% Celluclast® 1.5 L + 0.5% Pectinex® XXL + 0.4 M 甘露醇,培养 8 小时,从叶肉组织中分离出原生质体。利用藻酸盐薄层(TAL)方法包埋原生质体。随后,在添加了 3%蔗糖、2,4-二氯苯氧乙酸(2,4-D,0, 0.56, 1.12 µM)和植物生长调节剂(PSK,0, 0.05, 0.1 µM)的 Murashige and Skoog(MS)培养基中培养 TAL 片段。利用响应面方法(RSM)和中心复合设计(CCD)预测了 2,4-D 和 PSK 引发原生质体分裂、多细胞发育和微胼胝体形成的最佳浓度。根据 CCD 反应,确定 1.12 µM 2,4-D 和 0.065 µM PSK 是诱导原生质体分裂和微茧形成的最佳浓度。所获得的结果对于大花罂粟原生质体的分离和培养很有价值;然而,利用原生质体产生的胼胝体进行植物再生还需要协调努力。
Isolation, culture of Platycodon grandiflorus protoplasts: factors affecting protoplast yield, cell division, and micro-callus formation
Plyatocodon grandiflorus is an important medicinal herb, its rhizomes are utilized as vegetables and as medicinal food. In this investigation, we were able to successfully isolate protoplasts from P. grandiflorus leaf mesophyll tissues in order to promote the development of calli. It has been determined what are the ideal conditions for protoplast isolation, involving the combination of enzymes, the osmoticum, and the length of enzyme digestion. For the most effective results, cell protoplast washing (CPW) medium with 1.0% Viscozyme® L + 3% Celluclast® 1.5 L + 0.5% Pectinex® XXL + 0.4 M mannitol and 8 h of incubation was used to isolate protoplasts from leaf mesophyll tissues. The thin alginate layer (TAL) approach was utilized to embed the protoplasts. Subsequently, TAL segments were cultured in Murashige and Skoog (MS) media supplemented with 3% sucrose, 2,4-dichlorophenoxyacetic acid (2,4-D, 0, 0.56, 1.12 µM), and phytosulfokine (PSK, 0, 0.05, 0.1 µM). The optimal concentration of 2,4-D and PSK for initiation of protoplast division, multicell development, and micro-callus formation was predicted using response surface methodology (RSM) with central composite design (CCD). Based on the CCD responses, it was determined that 1.12 µM 2,4-D and 0.065 µM PSK were the best concentrations to induce protoplast division and the formation of micro-calluses. The acquired results are valuable for P. grandiflorus protoplast isolation and culture; however, coordinated efforts are required for plant regeneration using callus produced from protoplasts.
期刊介绍:
Horticulture, Environment, and Biotechnology (HEB) is the official journal of the Korean Society for Horticultural Science, was launched in 1965 as the "Journal of Korean Society for Horticultural Science".
HEB is an international journal, published in English, bimonthly on the last day of even number months, and indexed in Biosys Preview, SCIE, and CABI.
The journal is devoted for the publication of original research papers and review articles related to vegetables, fruits, ornamental and herbal plants, and covers all aspects of physiology, molecular biology, biotechnology, protected cultivation, postharvest technology, and research in plants related to environment.