An efficient protocol for Ulmus minor Mill. Protoplast isolation and culture in agarose droplets

• Type: Article in International Scientific Journal
• Date: 2006

Authors

Conde, P (Author) Other; Santos, C (Author) Other

Journal

Title: Plant Cell, Tissue and Organ Culture

Vol. 86

Pages: 359-366

ISSN: 0167-6857

Publisher: Springer Nature

Indexing

ISI Web of Knowledge - 19 Citations

Scopus - 21 Citations

Other information

Authenticus ID: P-004-HQQ

DOI: 10.1007/s11240-006-9122-2

Abstract (EN): We describe here an efficient and reproducible protocol for isolation and culture of protoplasts from Ulmus minor. Different sources of donor tissues were tested for protoplast isolation: callus and juvenile leaves from in vitro and greenhouse plants. Several combinations and concentrations of hydrolytic enzymes were used. Comparative tests between Cellulase Onozuka R10 and Cellulase Onozuka RS were made and the last one proved to be more efficient. Both the pectinases used, Macerozyme Onozuka R10 and Pectinase (Sigma((R))), were efficient in protoplast isolation and there was no need for a more active pectinase. In vitro leaves proved to be the best source for protoplast isolation and produced an average of 3.96 x 10(7) protoplasts per gram of fresh weigh. Elm mesophyll protoplasts were cultured using the advantageous method of agarose droplets and a modification of the Kao and Michayluk culture medium, using two plating densities (1 x 10(5) and 2 x 10(5) protoplasts ml(-1)). Protoplast division and evolution into colonies and microcalli was promoted in the agarose droplets plated at 2 x 10(5) protoplasts ml(-1). Ten weeks after protoplast culture initiation a plating efficiency of 2.7% was attained and the bigger microcalli, with at least 0.5 mm diameter, were transferred to a solid medium previously used for the production of embryogenic callus.

Language: English

Type (Professor's evaluation): Scientific

No. of pages: 8