A Study to Develop an in Vitro Dual Culture System with Carrot Hairy Roots and Arbuscular Mycorrhizal Fungi

doi:10.3969/j.issn.1000-2561.2020.08.005

Abstract

Abstract:

Agrobacterium rhizogenes K599 was used to induce carrot hair roots. An in vitro dual culturing system with Funneliformis mosseae and carrot hairy roots was then developed, and factors including methods to obtain axenic, supersonic processing, chilling treatment and medium pH that might affect both AM fungal spores germination and dual culturing were optimized. It was found that the third axenic method resulted in the highest germination rate (48.88%) and the lowest contamination rate (9.98%) at 15 days. Supersonic processing helped reduce AM fungal spores’ contamination by 11.17% to 14.53%, while a chilling pretreatment at 4℃ for 10 days and 15 days increased the germination rate of AM fungal spores to 70% and 65%, respectively. In water agar, the highest germination rate was achieved at a pH of 6.5 (48.50%), whereas spore germination was inhibited when pH was lower than 5.5 or higher than 8.0, with germination rate decreased to 15.43% at pH 5.5 and 16.06% at pH 8.0. Furthermore, it was found that the success rate of dual culturing was enhanced when sterilized spores were first germinated on agar medium and long hyphae were then selected for inoculation with their extension direction towards the growth direction of the carrot root. The MSR medium was found to be optimal for dual culture. The successful in vitro dual culture of carrot hairy roots and arbuscula mycorrhizal fungi provides an ideal experimental system to obtain pure AM fungal spores and to conduct future studies on the physiological and molecular mechanism of carrot mycorrhiza.

Key words: carrot hair roots, Agrobacterium rhizogenes, arbuscular mycorrhizal fungi, Funneliformis mosseae, dual culture

CLC Number:

S154.3

ZHANG Jinlian,LIU Jinhua,BAO Han,LI Dongping,SONG Juan,HUANG Jinghua,CHEN Tingsu. A Study to Develop an in Vitro Dual Culture System with Carrot Hairy Roots and Arbuscular Mycorrhizal Fungi[J]. Chinese Journal of Tropical Crops, 2020, 41(8): 1535-1542.

Figures/Tables 7

References 34

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消毒方法 Disinfection methods	萌发率Germination rate/%			污染率Contamination rate/%
消毒方法 Disinfection methods	3 d	9 d	15 d	3 d	9 d	15 d
1	14.17±2.21^a	37.50±4.17^b	42.50±3.00^b	3.53±3.57^a	13.33±4.78^b	14.72±2.80^b
2	12.65±2.06^a	33.71±0.66^b	35.94±3.57^c	3.41±3.45^a	28.79±5.58^a	31.02±3.08^a
3	16.79±4.76^a	47.65±0.85^a	48.88±2.67^a	1.10±1.92^a	8.52±4.49^b	9.98±3.33^b

消毒方法 Disinfection methods	萌发率Germination rate/%			污染率Contamination rate/%
消毒方法 Disinfection methods	3 d	9 d	15 d	3 d	9 d	15 d
1	14.17±2.21^a	37.50±4.17^b	42.50±3.00^b	3.53±3.57^a	13.33±4.78^b	14.72±2.80^b
2	12.65±2.06^a	33.71±0.66^b	35.94±3.57^c	3.41±3.45^a	28.79±5.58^a	31.02±3.08^a
3	16.79±4.76^a	47.65±0.85^a	48.88±2.67^a	1.10±1.92^a	8.52±4.49^b	9.98±3.33^b

超声波 Supersonic	萌发率Germination rate/%			污染率Contamination rate/%
超声波 Supersonic	1	2	3	1	2	3
处理	42.50±3.00^a	35.94±3.57^a	48.89±2.67^a	14.71±2.80^a	31.02±3.83^a	9.63±2.23^a
不处理	43.09±3.22^a	36.92±3.30^a	48.76±1.07^a	25.88±5.91^b	45.55±5.09^b	23.06±5.56^b

超声波 Supersonic	萌发率Germination rate/%			污染率Contamination rate/%
超声波 Supersonic	1	2	3	1	2	3
处理	42.50±3.00^a	35.94±3.57^a	48.89±2.67^a	14.71±2.80^a	31.02±3.83^a	9.63±2.23^a
不处理	43.09±3.22^a	36.92±3.30^a	48.76±1.07^a	25.88±5.91^b	45.55±5.09^b	23.06±5.56^b

pH	萌发率Germination rate/%
pH	3 d	6 d	9 d	12 d	15 d	18 d
5.5	2.34±2.03^c	11.85±3.92^d	15.43±1.07^d	15.43±1.07^d	15.43±1.07^d	15.43±1.07^d
6.0	11.48±1.70^b	27.53±2.17^b	33.21±3.52^c	34.32±5.26^c	34.32±5.26^c	34.32±5.26^c
6.5	16.79±4.76^a	35.55±3.85^a	47.65±0.84^a	48.49±2.67^a	48.49±2.67^a	48.49±2.67^a
7.0	12.59±3.57^ab	33.33±3.35^a	41.48±4.20^b	41.48±4.20^b	41.48±4.20^b	41.48±4.20^b
7.5	5.80±2.17^c	19.63±2.89^c	27.78±5.09^c	27.78±5.09^c	27.78±5.09^c	27.78±5.09^c
8.0	1.10±1.91^c	11.92±2.45^d	16.06±2.48^d	16.06±2.48^d	16.06±2.48^d	16.06±2.48^d