1. Genetic Diversity in Pollen Abiotic Stress Tolerance John J. Burke USDA-ARS, Lubbock, TX

2. Cotton flowers bloom for one day. The petals open in the morning, and by evening the petals close and begin to senesce. Because of the brief window of time available for pollination to occur, cotton pollen must be ready to germinate rapidly upon contacting the stylar tissues. Cotton pollen is classified by plant systematists as spherical, polyporate, and partially hydrated. Being partially hydrated accelerates the germination process, but may predispose the pollen grain to increased sensitivity to temperature and relative humidity fluctuations.

3. This study evaluated mature pollen of six cotton lines with known differences in pollen development and dehiscence when exposed to high temperature stress in the field. We evaluated the in vitro temperature sensitivity of the pollen from non-stressed plants to determine if diversity in high temperature stress tolerance exists within these lines. Secondly, we evaluated the humidity sensitivity of the mature pollen from these lines.

4. The putative heat tolerant lines showed less sterility than the high fiber quality lines in both the laboratory and field evaluations. Line (Percy) ST 474 3.43 SG 248 3.70 DP 565 4.03 NM 67 4.25 Acala Maxxa 4.28 Phy. 72 4.75 Pollen rating scale: 1 = 100% fertility 2 = 75% fertility 3 = 50% fertility 4 = 25% fertility 5 = 0% fertility

5. In vitro analysis of the genetic diversity in the temperature sensitivity of mature cotton pollen was evaluated using the procedure of Burke et al. (Agron. J 96: 359-368. 2004.)

6. Flowers were incubated at either 28C or 35C under high humidity for 5 hours. Pollen was harvested and the percent germination in vitro evaluated after a 28C, 80%RH, 1 hour incubation.

7. How does humidity impact pollen germination and tube development?

8. Room temperature - low humidity 6.5 hour challenge followed by germination at 28C, 80%RH for 1 hour Control

9. Germination for 1 hour at 28C in either 25% or 80% relative humidity - greenhouse grown

11. Time: 0 sec Time: 45 sec Time: 60 sec Time: 75 sec Time: 90 sec Time: 105 sec Time: 120 sec Time: 135 sec Time Lapse of Pollen Rupturing in Water

12. Germination for 1 hour at 28C in either 25% or 80% relative humidity - greenhouse grown

13. Greenhouse Time to first pollen grain rupture in 0.8M sucrose - greenhouse grown

15. NM67 PHY72 3 Minutes in 0.8 M Sucrose

17. Rapidly dehydrating pollen reduces the risk of gene transfer. Slowly dehydrating pollen increases the probability of gene transfer for hybrid production.

19. Preliminary results suggest varietal differences in pollen germination and growth rate upon pollination.

20. We investigated mature pollen characteristics and identified distinct genetic variability in the low humidity stress tolerance, but no diversity in high temperature tolerance. We have developed laboratory-based tests permitting rapid evaluation of the overall abiotic stress tolerance of the pollen. We showed that the two most heat sensitive lines from field studies are the two lines most sensitive to low humidity. Our findings provide breeders with a previously unexplored reservoir of genetic diversity associated with reproductive abiotic stress tolerance.