| Elymus sibiricus L. is a kind of high-quality forage widely distributed in north and northwest alpine region of China. With important nutritional and ecological values, it is stored mainly as seed form at low-temperature germplasm library. However, it only prolongs the shelf-life of cryopreservation, aging could be caused by long-term storage. Seed aging will not only reduce the germination rate of germplasm, but also lead to physiological and biochemical characteristics changes, and genetic variations of protein and DNA might be aroused. The germplasm storage shall be updated after a certain time in order to maintain the genetic integrity.In this research Elymus sibiricus L. bred varieties (Elymus sibiricus L. cv Nongmu, Elymus sibiricus L. cv Chuancao No.1) and two wild materials (Elymus sibiricus L.01090, Elymus sibiricus L.03658) were selected. The seeds of them were treated in the artificial aging conditions at high-temperature (40?) and high humidity (70% RH) of aging method to obtain different germination rate (80%,50% and 30%) by taking the seeds with more than 90% germination rate with no aging treatment as a control. Elymus sibiricus L. were used to study effects of seed aging in the alteration of genetic integrity by gliadin and ISSR molecular markers. The purpose of this study was to provide foundation for making a criteria of preserving and regenerating germplasm resources to Elymus sibiricus L.. The main results were showed as follows:1.Four Elymus sibiricus L. materials with different germination rate presented the declined trend with the increase of aging time. The time was different when the aging of different materials got to the same germination rate. The time of two bred varieties required was longer than that of the wild materials. The aging time for the bred varieties was 4d,6d and 7d when germination rate dropped to about 80%,50% and 30%. However, the aging time for the wild materials 01090 was 4d,5d and 6d, while the aging time for the wild materials 03658 was 3d,4d and 5d.2.The results of effects on genetic integrity of different seed aging treatments to four Elymus sibiricus L. materials by gliadin markers showed that numbers of gliadin bands changed after aging mainly in areas of ? and ?. The genetic parameters of the germplasm population such as the number of alleles, the number of effective alleles, index of genetic diversity and Shannon's information index decreased compare to that of the control with the decreasing of germination rate. When germination rate dropped to about 30%, the number of alleles per loci of four materials showed significant differences compare to that of control. The index of genetic diversity of Elymus sibiricus L. cv Nongmu and Elymus sibiricus L. cv Chuancao No.1 showed significant differences compare to that of control, and the Shannon's information index of Elymus sibiricus L.01090 showed significant difference compare to that of the control.3.The results of effects on genetic integrity of different seed aging treatments to four Elymus sibiricus L. materials by ISSR markers showed that 108 bands were amplified by using 13 primers, of which 93 loci were polymorphic, and the average percentage of polymorphic loci was 86.09% for the test materials. Bands of 250bp-500bp and 750bp-2000bp appeared or disappeared after aging. The genetic parameters of the control such as the number of alleles, the number of effective alleles, index of genetic diversity, Shannon's information index and the percentage of polymorphic loci were higher than that of the aging populations. The genetic parameters of population reduced gradually with the increase of aging time. When the germination rate dropped to about 30%, the number of alleles per loci and the index of the genetic diversity of four materials showed significant differences compare to that of control. The germplasm of Elymus sibiricus L. should be suggested to update when the germination rate dropped to about 30%.4.Both gliadin markers and ISSR markers could be used for detecting effects on genetic integrity of seed aging for Elymus sibiricus L. germplasm resources. The results of the correlation analysis of genetic identity of the two markers showed highly significant positive correlation. The number of polymorphic loci detected by ISSR markers was more than that of polymorphic loci detected by gliadin markers. Therefore, ISSR markers are more applicable. |
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