Conservation Technology of In Vitro Tubers of Pinellia ternata by Delayed Growth

doi:10.7525/j.issn.1673-5102.2026.01.009

Abstract

Abstract:

To establish a technology system for the delayed growth preservation of Pinellia ternata tubers in vitro， this study valuated different sucrose mass concentrations， the ratio of sucrose to mannitol， and low-temperature methods to preserve the in vitro tubers. After 360 days， the tubers were allowed to resume growth， and the growth indicators were observed and statistically analyzed. Starch grain dynamics in the tubers were examined using paraffin sectioning technique. The genetic stability of regenerated plants after preservation was assessed using ISSR-PCR and RAPD molecular markers. Mature tubers were inoculated on fresh 1/2MS+30 g⋅L^-1 sucrose+60 g⋅L^-1 mannitol medium and stored at 4 ℃ for 360 days， the sprouting rate of tubers after recovery growth was as high as 96.67%， and the plants grew well with the best preservation effect. Tubers cultured on medium containing 90 g⋅L^-1 sucrose， showed reduced growth despite a sprouting rate of 90.00%， likely due to the excessively high osmotic pressure. When mature tubers were stored in the old medium of 1/2MS+30 g⋅L^-1 sucrose at 4 ℃ for 360 days， the sprouting rate of tubers after recovery growth was the lowest， only 36.70%. Histological studies showed that in the dead tubers after preservation， there were a few leaf primordia， almost no starch grains in the cells， and no mucilage cells. Both the high sprouting rate and low sprouting rate treatments had a large amount of starch grains stored in the tubers after recovery growth， but the treatment with high survival rate had more layers of leaf primordia in the stem tips and fewer mucilage cells， and the plants grew more vigorously. A total of 1 140 bands were amplified using molecular marker techniques， and no variant bands were detected， indicating genetic stability of regenerated plants after delayed growth preservation. This study established a simple and efficient technique for the delayed growth preservation of P. ternatain vitro tubers， providing a feasible approach for short-term preservation of P. ternata germplasm.

Key words: Pinellia ternata, in vitro tuber, delayed growth preservation, regenerated plants, starch granule distribution, genetic stability

CLC Number:

Q944.5

Yanhong ZHANG, Yaping ZHANG, Linjia WANG, Chunyu HE, Qingyi GUO. Conservation Technology of In Vitro Tubers of Pinellia ternata by Delayed Growth[J]. Bulletin of Botanical Research, 2026, 46(1): 101-110.

Figures/Tables 9

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引物 Primer	序列（5′→3′） Sequence（5′→3′）	退火温度 Annealing temperature/℃
Em2	GACTGCGTACGAATTTGC	53.0
Em13	GACTGCGTACGAATTCTG	50.5
Lect7	GTTGCTGCTCGAGAGACGA	57.9
881	GGGTGGGGTGGGGTG	60.6
835	AGAGAGAGAGAGAGAGYC	50.2
854	TCTCTCTCTCTCTCTCRG	56.0

保存时间 Preservation time/d	保存块茎数量 Number of preserved tubers	芽尖萌动数量 Number of bud tips sprouting	萌动率 Sprouting rate/%	生根数量 Number of rooting	生根率 Rooting rate/%	块茎状态 Status of tubers
40	30	0	0	0	0	饱满
80	30	3	10.0	0	0	饱满
120	30	4	13.3	2	6.7	饱满
160	30	8	26.7	5	16.7	略干缩
200	30	8	26.7	5	16.7	略干缩
240	30	8	26.7	5	16.7	略干缩
280	30	8	26.7	5	16.7	略干缩
320	30	8	26.7	5	16.7	略干缩
360	30	8	26.7	5	16.7	略干缩

保存时间 Preservation time/d	保存植株数量 Number of preserved plants	新鲜叶柄数量 Number of fresh petioles	干枯叶柄数量 Number of withered petioles	叶柄干枯率 Withered petiole rate/%
40	30	79	6	7.06
80	30	73	12	14.12
120	30	46	39	45.88
160	30	21	64	75.29
200	30	0	85	100.00
240	30	0	85	100.00
280	30	0	85	100.00
320	30	0	85	100.00
360	30	0	85	100.00

高渗透压处理 High osmotic pressure treatment	保存块茎数量 Number of preserved tubers	恢复培养后成苗率 Seedling rate after recovery culture/%	每块茎的叶柄数量 Number of petioles per tuber	叶色 Leaf color	平均叶柄长度 Average length of petiole/cm
30 g·L^-1蔗糖	30	76.67^c	2.81^bc	绿色	4.75^ab
60 g·L^-1蔗糖	30	76.67^bc	2.87^b	绿色	3.72^cd
90 g·L^-1蔗糖	30	90.00^ab	3.05^ab	绿色	3.05^d
30 g·L^-1蔗糖+20 g·L^-1甘露醇	30	70.00^cd	3.46^a	绿色	4.21^bc
30 g·L^-1蔗糖+40 g·L^-1甘露醇	30	73.33^cd	3.10^ab	绿色	3.36^cd
30 g·L^-1蔗糖+60 g·L^-1甘露醇	30	96.67^a	3.44^a	绿色	4.24^bc
60 g·L^-1蔗糖+20 g·L^-1甘露醇	30	60.00^cd	2.32^c	绿色	3.52^cd
90 g·L^-1蔗糖+20 g·L^-1甘露醇	30	66.67^cd	3.50^a	绿色	5.47^ab