Cryoconservation of in vitro grown shoot tips of Cicer microphyllum: A crop wild relative of chickpea.

IF 1 4区生物学 Q3 BIOLOGY Cryo letters Pub Date : 2023-11-01

S Chander, R Gowthami, R Pandey, M Shankar, A Agrawal

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引用次数: 0

Abstract

Background: Cicer microphyllum Benth. is a crop wild relative (CWR) of chickpea (C. arietinum L.), that possess useful genes for cold and drought tolerance. The species is being conserved in the In Vitro Active Genebank for short- to medium-term conservation. Cryopreservation would be a useful complementary approach for its long-term conservation.

Objective: The current work aimed to develop an efficient cryoconservation protocol for cryobanking of C. microphyllum shoot tips.

Materials and methods: In vitro shoot tips excised from 4-month old shoot cultures grown on B5 + 0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO₃ medium were cryoconserved using a droplet-vitrification technique. Post-thaw regrowth was evaluated after: (i) preculture medium (B5 basal, B5 + 3, 4, 6 and 10% sucrose), (ii) preculture incubation temperature (25 ± 2, 10, 8 and 22/5 degree C), (iii) PVS2 duration (10, 20, 30. 40, 50 and 60 min) and (iv) regrowth medium (B5) supplemented with 0.5 mg/L KIN + 0.1 NAA mg/L; 0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO₃; 0.2 mg/L BAP + 10 mg/L AgNO₃; 0.2 mg/L BAP + 20 mg/L AgNO₃ and 0.2 mg/L BAP + 30 mg/L AgNO₃.

Results: In vitro shoot tips grown on B5 + 0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO₃, precultured on B5 + 6% sucrose at 10 degree C for 3 days, followed by PVS2 treatment for 20 min, unloading solution for 60 min and regrowth on B5 + 0.2 mg/L BAP + 20 mg/L AgNO₃ resulted in highest survival (57%) and regrowth (40%) after cryoconservation.

Conclusion: The standardized protocol was successfully used for cryobanking of in vitro shoot tips of C. microphyllum in the In Vitro Base Genebank of ICAR-NBPGR, New Delhi. Doi.org/10.54680/fr23610110412.

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鹰嘴豆野生近缘作物 Cicer microphyllum 离体生长嫩枝尖的低温保存。

背景：Cicer microphyllum Benth.是鹰嘴豆（C. arietinum L.）的一种作物野生近缘种（CWR），具有耐寒和耐旱的有用基因。该物种目前保存在体外活性基因库中，用于中短期保存。低温保存将是长期保护该物种的有效补充方法：目前的工作旨在开发一种有效的冷冻保存方案，用于冷冻保存 C. microphyllum 芽尖：从 B5 + 0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO3 培养基上生长 4 个月的离体芽尖上切除的芽尖采用液滴-玻璃化技术进行低温保存。在下列情况下对解冻后的再生进行了评估：(i) 预培养基（B5 基础培养基、B5 + 3、4、6 和 10%蔗糖）；(ii) 预培养孵育温度（25 ± 2、10、8 和 22/5 摄氏度）；(iii) PVS2 持续时间（10、20、30、40、50 和 60 分钟）；以及(iv) PVS2 的温度。40、50 和 60 分钟）；(iv) 添加 0.5 mg/L KIN + 0.1 mg/L NAA、0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO3、0.2 mg/L BAP + 10 mg/L AgNO3、0.2 mg/L BAP + 20 mg/L AgNO3 和 0.2 mg/L BAP + 30 mg/L AgNO3 的再生培养基（B5）：在B5 + 0.5 mg/L KIN + 0.1 mg/L NAA + 10 mg/L AgNO3上生长的离体芽尖，在B5 + 6%蔗糖上10摄氏度预培养3天，然后PVS2处理20分钟，卸载溶液60分钟，在B5 + 0.2 mg/L BAP + 20 mg/L AgNO3上重新生长，结果低温保存后的存活率（57%）和重新生长率（40%）最高：该标准化方案被成功地用于冷藏新德里ICAR-NBPGR体外基地基因库中的C. microphyllum体外芽尖。Doi.org/10.54680/fr23610110412.

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来源期刊

Cryo letters 生物-生理学

CiteScore

1.80

自引率

10.00%

发文量

审稿时长

1 months

期刊介绍： A bimonthly international journal for low temperature sciences, including cryobiology, cryopreservation or vitrification of cells and tissues, chemical and physical aspects of freezing and drying, and studies involving ecology of cold environments, and cold adaptation The journal publishes original research reports, authoritative reviews, technical developments and commissioned book reviews of studies of the effects produced by low temperatures on a wide variety of scientific and technical processes, or those involving low temperature techniques in the investigation of physical, chemical, biological and ecological problems.