Sukhraj S. Dhillon

DNA, RNA, protein and heterochromatin changes during embryo development and germination of soybean (Glycine max L.)

SummaryDNA, RNA, protein and heterochromatin were measured cytophotometrically in developing soybean (Glycine max) seeds. The average 2C DNA content for the soybean genome was 2.64 pg. The amounts of nuclear DNA in embryo axes showed no significant change during embryo development, whereas the DNA content in cotyledon nuclei increased significantly from 3.58 pg to 5.49 pg. The number of endopolyploid nuclei increased from 26% to 48% and the DNA content from 4.45 to 5.49 pg after cessation of cell division. The changes in RNA and protein content during embryo development were in general similar to those in DNA content. This can be interpreted that increased DNA levels in soybean cotyledons generated during embryogeny increase the protein synthesizing capacity. During the first 15 days of germination, the number of endopolyploid nuclei in cotyledons declined from 46% to 4%, and this decline is interpreted as DNA degradation providing a ready source of nucleosides and phosphates during early embryo growth. A later decline, however, between 15 and 20 days after germination, was age related similar to leaf senescence, because the percentage of endopolyploid nuclei remained unchanged while the number of non-viable cells increased. In senescing cotyledons, 73% and 80% of RNA and protein but only 20% of DNA were lost, as compared to dormant cotyledons. The heterochromatin (condensed chromatin) measurements indicated that nuclei of metabolically inactive dormant and senescent cotyledon nuclei contained an average of 33% more heterochromatin than nuclei from the green cotyledons of seedlings.

Feulgen cytophotometry of pine nuclei. II : Effect of pectinase used in cell separation

Pectinase used for cell separation prior to cytophotometry contains a DNase that is able to penetrate the cells of pine root tips and attack nuclear DNA. When pine root tips were exposed to 1% pectinase (pH 6.0), there was a decrease in nuclear DNA content at every sample point and a sharp drop between 16 and 20 hr. The effect of the DNase was eliminated by preparing the enzyme solution in 0.01 M sodium citrate or 0.001 M EDTA. It is suggested that heat denaturation of the DNase should also be effective and might be used in combination with the magnesium chelators.

DNA Analysis During Growth and Development

Nuclear DNA changes in relation to growth and development were followed in Populus deltoides. These observations were supported with findings in other plant species. The genome size of P. deltoides was small and was comprised of a 2C DNA content of 1.07 pg. The genome showed an average increase of 27% under long-day photoperiod as compared to short-day photoperiod. Plants under long-day conditions displayed significantly higher growth. Furthermore, the young leaves possessed higher DNA values as compared to mature leaves. In view of our present as well as previous observations in several species, it appears that higher DNA values are related to increasing protein synthesis for maintaining growth. The senescing leaves displayed an average reduction in nuclear DNA content of 21% as compared to mature leaves. Purified DNAs from nuclei and chloroplasts of mature and senescing leaves did not display different thermal profiles. However, chloroplast DNA displayed a significantly higher %GC ratio than nuclear DNA in mature leaves. The %GC was 33.92 for nuclear and 35.95 for chloroplast DNA. Studies are in progress to understand the growth- and senescence-related DNA changes at the molecular level.