# An Efficient Micropropagation Protocol for Camellia chekiangoleosa ‘Ganhongyou 1’ via Stem Segment Culture

## TL;DR

This paper develops an efficient method to propagate the Camellia chekiangoleosa ‘Ganhongyou 1’ plant using stem segment culture, achieving high survival and growth rates.

## Contribution

A novel micropropagation protocol for Camellia chekiangoleosa ‘Ganhongyou 1’ with optimized growth regulator combinations and a phloem-rooting pattern confirmation.

## Key findings

- Optimal bud induction achieved with 86.67% success using 6-BA and IAA on Hyponex medium.
- A combined in vitro and ex vitro strategy resulted in an 88% survival rate after rooting and acclimatization.
- Adventitious roots originated from phloem parenchyma cells, confirming a phloem-rooting pattern.

## Abstract

The provincial-level registered superior cultivar Camellia chekiangoleosa ‘Ganhongyou 1’ boasts superior economic traits coupled with significant ornamental value, driving demand for an efficient propagation system. Consequently, this study aimed to develop a rapid micropropagation protocol by investigating culture conditions using semi-woody nodal segments with axillary buds as explants on Hyponex basal medium supplemented with varying combinations of plant growth regulators. Contamination was effectively minimized to 18% by a combined approach of surface sterilization (75% ethanol, 0.1% HgCl2, and 20% NaClO) and incorporating 1 mL/L bactericide into the induction medium. For bud induction, the optimal medium was 2 g/L Hyponex supplemented with 1.0 mg/L 6-benzylaminopurine (6-BA) and 0.2 mg/L indole-3-acetic acid (IAA), achieving an 86.67% induction rate. The best proliferation was achieved on the medium containing 2 g/L Hyponex, 1.0 mg/L 6-BA, 0.15 mg/L 3-indolebutyric acid (IBA), and 0.5 mg/L gibberellic acid (GA3), yielding a proliferation coefficient of 6.53. A combined strategy, integrating in vitro pre-culture with ex vitro treatment, proved most effective for rooting and acclimatization: shoots were first pre-cultured for 20 days on 1/2 strength Hyponex medium supplemented with 0.5 mg/L 1-naphthaleneacetic acid (NAA) and 2.0 mg/L IBA, followed by ex vitro base treatment with 1.0 g/L ABT (a rooting powder complex) solution before transplantation into seedling bags. This approach resulted in an 88% survival rate. Furthermore, anatomical analysis revealed the origin of adventitious root primordia from phloem parenchyma cells, thereby confirming a phloem-rooting pattern for this species. In conclusion, this study establishes a practical and efficient micropropagation protocol for ‘Ganhongyou 1’, providing a reliable technical foundation for its commercial-scale seedling production.