Cytological analysis of plants regenerated from maize tissue cultures
Plants may be regenerated from maize callus tissue growing on an appropriate medium (Green and Phillips, 1975, Crop Sci. 15:417). Plant tissue cultures often possess aneuploid and polyploid cells. The data reported here give a preliminary indication as to whether or not the chromosome constitution and behavior of plants regenerated from maize tissue cultures, derived according to the methods described in the above reference, are normal. The cultures used in this study were initiated from A188 x W22 A C R-nj b pl embryos 1.5-2.0 mm in length (12 days postpollination) on MS medium containing 1 mg 2,4-dichlorophenoxyacetic acid (2,4-D) per liter. Plants were regenerated during the fourth through sixth subcultures, 65-125 days after culture initiation, by transferring callus to asparagine-minus MS medium which contained 0 or 0.25 mg 2,4-D/l. One hundred eight plants collected during this period were transplanted to vermiculite, watered with a 1/4 strength MS salts solution and allowed to grow for 10-14 days in an incubator at 70-75% relative humidity, 27-28 C, and a 16 hour photoperiod with a light intensity of 3500 lux. Eighty-seven plants (80%) survived and were transplanted to soil and grown to maturity in the greenhouse. At the appropriate stage, a portion of the immature tassel was collected from 43 plants and fixed in 3 parts 95% ethanol:1 part glacial acetic acid. The portion of the tassel not collected allowed for subsequent pollen sterility determinations on each plant.
Normal chromosome numbers, pairing, and pollen fertility levels were observed for 41 plants. The two cytologically aberrant plants were both mosaics. One had tetraploid tassel sectors while the other had aneuploid sectors which were observed to be monosomic for chromosome 5. Both plants possessed the chromosome pairing and behavior patterns expected with their respective changes in chromosome number. Pollen fertility was determined to be normal for both plants. Since the pollen analysis was based on only one sample per plant and performed prior to the cytological analysis, we assume that the expected pollen sterility was missed due to the sampling procedure. In these materials, therefore, 95% of the regenerated plants appeared to be cytologically normal. The aberrant types were mosaics. At what point in development the mosaics arose is unknown.
In addition to the above materials, eleven plants were regenerated from three-year-old maize callus tissue of the same genotype and by the same procedures as described above. These plants possessed oppositely arranged (decussate) leaves and were about three feet tall at maturity. Three were analyzed at pachynema and all possessed a heteromorphic chromosome 6 bivalent. One chromosome 6 in each plant was deficient for the distal one-third of the long arm. Since all three plants were cytologically identical, the callus tissue from which the plants were derived may have been homogeneous for the deficiency.
C. E. Green, R. L. Phillips and A. S. Wang
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