Transfer of foreign genes into intact maize cells with high-velocity microprojectiles

Abstract

This report describes a process for delivering foreign genes into maize cells that does not require the removal of cell walls and is capable of delivering DNA into embryogenic and nonembryogenic tissues. Plasmid harboring a chimeric chloramphenicol acetyltransferase (CAT) gene was adsorbed to the surface of microscopic tungsten particles (microprojectiles). These microprojectiles were then accelerated to velocities sufficient for penetrating the cell walls and membranes ofmaize cells in suspension culture. High levels of CAT activity were consistently observed after bombardment of cell cultures of the cultivar Black Mexican Sweet, which were comparable to CAT levels observed after electroporation of protoplasts. Measurable increases in CAT levels were also observed in two embryogenic cell lines after bombardment. Gene expression was observed only when an intron from the alcohol dehydrogenase 1 gene of maize was ligated between the 35S promoter and the CAT coding region. CAT activity was detected in cell cultures bombarded with microprojectiles with an average diameter of 1.2 ,gm, but not after bombardment with microprojectiles 0.6 or 2.4 ,Im in diameter. Bombarding the same sample several times was found to markedly enhance CAT activity. These results demonstrate that the particle bombardment process can be used to deliver foreign DNA into intact cells of maize. Because this process circumvents the difficulties associated with regenerating whole plants from protoplasts, the particle bombardment process may provide significant advantages over existing DNA delivery methods for the production of transgenic maize plants. In addition, the process should be of value for studying transient and stable gene expression within intact cells and tissues.

The moss Physcomitrella patens: methods and tools from cultivation to targeted analysis of gene function

“…The first technique especially developed for the transformation of plant cells and that is also applicable for the transformation of P. patens, is the particle or gene gun technology that delivers DNA biolistically directly into diverse plant tissues (Klein et al, 1988). The biolistic transformation method is not restricted to specific plants or tissue types and functions with the same efficiency in monocotyledonous and dicotyledonous plants (Sawahel et al, 1992).…”

Genetic Transformation of Maize Cells by Particle Bombardment

“…Previous work based on the analysis of transient gene expression has shown that microprojectiles can be used for the delivery of DNA to a wide range of intact plant cells and tissues including those from onion, maize, tobacco (13)(14)(15)(16), rice, wheat, and soybean (22). Stable transformation by particle bombardment has been shown in tobacco (16) and soybean (18).…”

Transformation of potato via Agrobacterium coated microparticle bombardment

“…Biolistic gene transfer, a method by which foreign genes may be introduced into cells via high velocity microprojectiles (Klein et al 1988) has been used in transforming monocots such as orchids (Suwanaketchanatit et al 2007) and dicots such as soybean (El-Shemy et al 2007), but has only recently been used in potato. Up to 0.5 transformants per leaf shot were achieved using particle bombardment (Craig et al 2005), almost two orders of magnitude higher than the first report in potato .…”