The effect of 8 days of microgravity on regeneration of intact plants from protoplasts

Abstract

T,-H, 1994. The effect of H days oi niicroEravily on receneralion of inuicl plants (rom protoplasts, -Phvsiol. Plant, 92; 404^1 I,"The growih and development of protoplasts of rapeseed {Brii.\.\iia napu\ L. cv Line) and carrol (Daucus carora L, cv. Na\ona> were studied onboard the Space ShuUle 'Discovery' during an 8-da\ Internationa] Microgravity Laboraloiy (IML-1) mission in January !992. The flight e\pei'imenls were carried out in 'Biorack'. a fully controlled cell biological experimental facilit\. under microgravity conditions and in a l-;i,' centrifuge. Parallel experiments were performed in a 'Biorack' m(tdule on the ground. After retrieval, some samples \^ele subcultured on appropriate media and analysed for callus growih and regeneration to inlact plants. The remainder v\ere used for biochemical analysis. Samples fixed on board the Space Shuttle were kepi in 1 ^. f glutaraldehyde fixative al 4°C for ^-^ days for microscopy analysis after retrieval. Prcttctplasls expctsed to micrograviiy conditiitns showed u delay in cell wall synthesis. Cells were swctllen in appearance and fttrmed cell aggregates wilh (tnlv tew cells. Callus were obtained from protoplasts cultured under niicrograviu (FOg), (tn ihe !-.£,' cenlrifuge on board the shultle (Fig), under normal l-,i.' c(tndilions <in the ground (Gig), and on a centrifuge on Ibe ground giving 1.4 x' (G1.4g), Regeneration of inlacl rapeseed planis was obtained from Fig. Gig and G1.4g. H(twever. no plants were regenerated from protoplasts exposed to microgravity iFOgi, Biochemical analysis indicaled thai ihe niicrogra\ily samples (FOg) displayed a reduced packed cell volume, an increased concentration of soluble proteins per cell, and a reduced specific acli\ ity of peroxidase in Ihe cyt(tplasm. Morphometric analysis of fixed samples demonstrated that .^-dav old proioplasls under micrograviu conditions were significantly larger than proutplasls kept on the l-j,' centrifuge in space. LItrastriictural analysis by transmission electron microscopy sh(twed that protoplasts exposed (o microgravity conditions for .i days bad larger vacuoles and a slightlv reduced starch content compared to Fig cells. Cell aggregates formed under microgravity conditions (FOg) had an average o! 2^ cells per aggregate while aggregates loniied under Fig had 8-12 cells.

Simulated weightlessness and hyper‐g results in opposite effects on the regeneration of the cortical microtubule array in protoplasts from Brassica napus hypocotyls

“…The increased size found in protoplasts exposed to simulated weightlessness on the clinostat (Fig. 6) is consistent with the observations done by astronaut Roberta Bondar during the IML-1 mission (Rasmussen et al 1994a) and the statistical analyses performed on cells after the IML-1 mis-sion (Rasmussen et al 1994b). The same effect was observed after the S/MM-03 mission in 1996 (Iversen et al 1999).…”

“…The results from the present experiments indicate that the reduced synthesis of cellulosic compounds and increased volume of the protoplasts exposed to micro‐ g conditions (Rasmussen et al 1992, 1994a,b; Iversen et al 1999) are correlated with effects on the CMTs in the protoplasts. The reduced CMT quantity may be an explanation for the increased cell size found in the present experiment and in the space experiments described, possibly by causing a low cellulose synthesis and/or an abnormal wall structure.…”

“…It should be noted that 90% of the 0 g‐ protoplasts died after 10 days. It is unlikely that the protoplasts inability to survive after rotation on the clinostat was caused by the loss of a g‐ vector since protoplasts cultivated under real micro ‐g conditions were able to regenerate roots or shoots after retrieval (Rasmussen et al 1994b).…”

“…Space‐flight experiments have indicated that protoplasts have a reduced production of cellulosic compounds and a decreased frequency of cell divisions under the conditions of weightlessness. In addition, protoplast volume has been observed to increase under micro ‐g conditions (Iversen et al 1992; Klimchuk et al 1992; Rasmussen et al 1992, 1994a,b; Iversen et al 1999).…”

The Utilization of Plant Facilities on the International Space Station—The Composition, Growth, and Development of Plant Cell Walls under Microgravity Conditions

“…The Biokosmos 9 experiment was followed by an experiment called “PROTO” which was flown on the Space Shuttle in 1992 with the IML-1 (International Microgravity Laboratory) mission [ 35 , 36 , 37 ]. Four years later, a new mission (S/MM-03; Shuttle Mission to Mir-03) was launched where a protoplast regeneration experiment was included.…”