Interaction of Culture Vessel Size, Medium Volume, and Carbon Dioxide Levels on the Growth of Various Plants in Vitro

Ultra-high levels of CO2, i.e., >10,000 ppm, enhance tissue culture growth and offers a relatively simple and inexpensive method to improve plant productivity in vitro. Growth responses employing ultra-high CO2 levels differ considerably in the literature. Unfortunately, various culture vessels and systems have been employed, making comparisons difficult. In this study, the influence of the vessel container size, medium volume, and various CO2 concentrations (0 to 50,000 ppm) was studied on the growth obtained from lettuce and spearmint cultures. All three of these factors influence growth responses from plants cultured in vitro. Vessel types tested included: culture tubes, Magenta containers, 1-quart jars, 0.5-gallon jars, and 1-gallon jars having culture volumes of 55, 365, 925, 1850, and 3700 ml, respectively. Increasing the size of the culture vessel resulted in an increase growth regardless of the CO2 level tested. For example, fresh weight of spearmint increases of >250% can be obtained in by employing a 1-quart jar compared to using a culture tube. Increasing medium volume using various vessel types, especially using high concentrations of CO2, resulted in dramatic growth increases. For example, a >100% increase in fresh weight could be obtained by increasing the medium volume from 50 ml to 100 ml within a 1-quart jar. These studies suggest that plant growth promoted by supplemental CO2 is limited by the culture vessel size and medium volume. Differences in growth responses obtained in past CO2 studies could be related to vessel type and medium volume as well as the CO2 levels employed. Future in vitro studies should consider these factors in the evaluation of the influence of Ultra-high CO2 levels on plant growth. Peculiar growth responses, especially pertaining to rooting and shooting exhibited by cultures grown in ultra-high CO2 levels will also be discussed.