Soybean protoplast culture and direct gene uptake and expression by cultured soybean protoplasts
A method was developed for culturing protoplasts freshly isolated from developing soybean (Glycine max L.) cotyledons. First cell divisions were observed within 5 days after protoplast isolation and microcalli, consisting of about 20 cells, were formed within 10 days. Thirty days after protoplast is...
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Published in | Plant physiology (Bethesda) Vol. 84; no. 3; pp. 856 - 861 |
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Main Authors | , , |
Format | Journal Article |
Language | English |
Published |
Rockville, MD
American Society of Plant Physiologists
01.07.1987
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Subjects | |
Online Access | Get full text |
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Summary: | A method was developed for culturing protoplasts freshly isolated from developing soybean (Glycine max L.) cotyledons. First cell divisions were observed within 5 days after protoplast isolation and microcalli, consisting of about 20 cells, were formed within 10 days. Thirty days after protoplast isolation, callus tissues were observed without the aid of a microscope. A 30 to 50% plating efficiency was consistently obtained. Using a polyethylene glycol-electroporation technique, DNA was introduced into these protoplasts. The protoplasts were then cultured to form callus. Chloramphenicol acetyltransferase (CAT) activity was detected in protoplast cultures 6 hours after introduction of a 35S-CAT-nopaline synthase 3′ chimeric gene. The highest CAT activity was detected in 3-day-old electroporated protoplast cultures, indicating transient expression of the introduced gene. Some CAT activity was detected in 40-day-old callus cultures and in geneticin (G418) selected callus tissues which also received a chimeric neomycin phosphotransferase II gene, indicating the presence of stable transformants. A control chimeric gene with an inverted 35S promoter failed to produce any CAT activity in this system. |
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Bibliography: | F60 F30 881958688 ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 0032-0889 1532-2548 |
DOI: | 10.1104/pp.84.3.856 |