Authora: Kopp, Richard F.; Maynard, Charles A.; Rocha de Niella, Patricia; Smart, Lawrence B.; Abrahamson, Lawrence P.
Author Address: College of Environmental Science and Forestry, Faculty of Environmental and Forest Biology, State University of New York, Syracuse, NY, 13210, USACollege of Environmental Science and Forestry, Faculty of Environmental and Forest Biology, State University of New York, Syracuse, NY, 13210.
Title: Collection and storage of pollen from Salix (Salicaceae).
Published in: American Journal of Botany, volumn 89, pages 248-252,(2002).
Abstract: " Genetic improvement of willows through traditional breeding can be facilitated by pollen collection and storage so that female flower receptivity need not be synchronized with pollen shed for breeding. Two experiments were completed to test the effectiveness of various organic solvents for willow pollen collection. In the first experiment, seven pollen collection treatments and an untreated control were tested with two willow clones. The other experiment tested three treatments that showed promise in the initial experiment and an untreated control with eight willow clones. Toluene and carbon tetrachloride were effective for pollen extraction, with average pollen germination percentages that were >15%, but both chemicals reduced pollen viability by 10-20% compared with an untreated control based on in vitro germination tests. Pollen extracted with carbon tetrachloride or toluene was successfully used in controlled pollination, and >100 new families were produced with this technique. Pollen viability remained high after 18 mo of storage at -20degreeC. Based on our results, toluene is the preferred solvent for future willow pollen extractions because it is as effective as carbon tetrachloride, is not a known carcinogen, and is less expensive."
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Authors: WAKI K; TAKEUCHI K
Authors Address: FAC. AGRIC., TAMAGAWA UNIV., TAMAGAWAGAKUEN 6-1-1, MACHIDA-SHI, TOKYO 194, JPN.
Title: POLLEN COLLECTING METHOD FOR ARTIFICIAL KIWIFRUIT POLLINATION 1. WASHING METHOD BY ORGANIC SOLVENTS

Published in: Bulletin of the Faculty of Agriculture Tamagawa University, volumn 30, pages 59-72, (1990).

Abstract: " Pollen collecting method for artificial kiwifruit pollination: 1. Washing method by organic solvents.To develop a labor-saving method for collecting pollen from Kiwifruit flowers, several procedures were examined; the following results were obtained. Pollen from pollen loads collected by honeybees showed a germination rate of 85%, but it was decreased to 40% after 4 days storage at a room temperature. When pollen loads were washed with successively decreasing concentrations of sucrose and finally with water, the germination rate was 78%. When pollen loads and pollen collected directly from flowers were washed with either 50%, 30% or 15% sucrose solution and then with water, their germination rates after drying were about 20%. This result suggests that these washing and drying methods are not suitable. The following 11 organic solvents out of 26 solvents used for washing showed high germination rates (over 50% of non-washed pollen): cyclohexane, n-pentane, isooctane, toluene, benzene, xylene, methyl acetate, ethyl acetate, diethyl ether, n-butyl alcohol and isoamyl alcohol. Pollen from early-stage male flowers did not germinate at all. But when it was washed with organic solvents after drying for 24 hours, its germination rate was as high as 77%. After 2 years of storing benzene-washed pollen at -17.degree. C, the germination rate was as high as 89%. Even when pollen was stored for 4 years below -17.degree. C, the germination ability was maintained to some extent (41% .apprx. 69%) when the pollen was washed with several solvents just after collection. Washing flowers with toluene, sucrose solution or water yielded more pollen than sifting although the latter two solvents reduced the germination rate considerably after drying. When toluene or benzene-washed pollen was used for hand pollination in the field, the percentage of fruit that set was high with no significant difference compared to non-washed pollen. The fruits obtained did not show any deterioration. Toluene and benzene are useful for obtaining pollen for artificial pollination."

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Title: STORAGE OF POLLEN GRAINS IN ORGANIC SOLVENTS EFFECTS OF SOLVENTS ON POLLEN VIABILITY AND MEMBRANE INTEGRITY
Authors: JAIN A; SHIVANNA K R
Authors Address: DEP. BOTANY, UNIV. DELHI, DELHI 110007, INDIA.
Published in: Journal of Plant Physiology, volumn 132, page 499-501,(1988).
Abstract: "Storage of pollen grains in organic solvents: Effects of solvents on pollen viability and membrane integrity.In vitro germinability was studied in pollen grains of Crotalaria saltiana Andr. stored in various organic solvents and correlated with pollen membrane integrity (as assessed by the fluorochromatic reaction test) and its permeability (as assessec by leaching of water soluble UV-absorbing materials into organic solvents during storage). Pollen grains stored in organic solvents with low dielectric constants (a measure of their non-polar nature) such as n-hexane, cyclohexane and diethylether showed high scores for germinability and membrane integrity, and very little leaching of UV-absorbing materials. Pollen grains stored in solvents with high dielectric constants (a measure of their polar nature) showed low scores for germinability and membrane integrity, and extensive leaching of UV-absorbing materials. The results indicate that viability of pollen grains stored in organic solvents is primarily determined by the effect of organic solvents on the integrity of pollen membrane."
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Title: Effect of storage in organic solvents on the germination of grapevine pollen
Published in: Journal of Horticultural Science, volumn 58, pages 389-392, (1983).
Author: P. K. AGARWAL
Author affiliation: Indian Institute of Horticultural Research, 255, Upper Palace Orchards, Bangalore-560080, India
Abstract: "Effect of storage in organic solvents on the germination of grapevine pollen. Grapevine pollen stored in amyl alcohol and benzene showed maximum germination even after 6 months storage, pollen tube growth being positively correlated with germination percentage."
The following is part of the introduction: "FOR grapevine breeding pollen storage is an important consideration, since different cultivars flower at different times and some may produce pollen when others have ceased flowering. Studies on the effects of varying humidity and temperature on pollen viability have been conducted (Gollmick, 1942; Nagarajan, et al., 1965; Randhawa et al., 1982) but there is no available information on the effect of organic solvents on pollen viability in any fruit crop. Preliminary studies on this technique of pollen storage in Camelia, Lilium and Impatiens spp. by Iwanami (1972) and Iwanami and Nakamura (1972) have suggested its usefulness for fruit crops.
This paper describes the effects of storage in ten organic solvents on pollen germinability in Vitis vinifera cv Anab-e-Shahi."
The following are some of the data that were given in Table 1. For each solvent three numbers are given. The first is the percent of germinated pollen, the second is the mean pollen tube length in microns, and the third is the correlation coefficient between the percent germination and the mean tube length.

"Caption of Table 1. Effect of 6 months storage in organic solvents on cv Anab-e-Shahi pollen germination and pollen tube growth
Control 5.33; 3.93; +0.84
Acetone 5.02; 10.03; +0,77
Amyl alcohol 57.59; 72.51; +0.48
Benzene 50.71; 59.92; +0.85
n-Butanol 42.80; 40.07; +0.91
Carbon tetrachloride 41.96; 25.37; +0.84
Ethyl acetate 38.54; 34.73; +0.74
Isoamyl acetate 52.36; 42.94; +0 57
Petroleum ether 41.92; 39.64; +0.79
Toluene (S-free) 37.37; 16.97; +0.67
Xylene 43.86; 49.93; +0.76"

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Title: Studies on the artificial pollination of watermelon. 1. On the storage of pollens of watermelon by the utilization of organic solvents.
Authors: Shimizu, Tatsuo. Tottori
Authors affiliation: Veg. Ornamental Crops Exp. Stn., Tottori, Japan.
Published in: Tottori-ken Yasai Shikenjo Kenkyu Hokoku, volumn 4, pages 9-17, (1983).
Abstract: "Utilization of org. solvents for the storage of watermelon pollens was examd. with ref. to the extensions of their pollen storage period. Among 6 org. solvents tested for pollen storage, ethyl acetate [141-78-6] and ethyl ether [60-29-7] were suitable. The effective storage period for pollens in Et2O was extended by drying the pollen after ether extn. from anthers. Na2SO4 addn. to Et2O and low temp. during storage were effective for extension of the pollen storage period. After removal of Et2O or EtOAc, .apprx.50% of the pollen lost its germination ability within 2 h at room temp. Thus, pollination with stored pollens must be completed immediately after the removal of the solvents. Satisfactory fructification and fruit-thickening were obtained by actual crossing with the pollen following storage in Et2O and EtOAc for 2 wk. For the practical application, the crossing at lower temp. period should be examd."

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Title: STORAGE OF POLLEN GRAINS OF CROTALARIA-RETUSA IN OILS
Authors: JAIN A, SHIVANNA K R

Published in: Sexual Plant Reproduction, Volumn 3, pages 225-227, (1990).

Abstract: "Storage of pollen grains of Crotalaria retusa in oils. Attempts were made to store pollen grains of Crotalaria retusa L. in a mineral oil (paraffin oil) and two vegetable oils (soybean oil and olive oil). Under laboratory conditions pollen grains not stored in oil lost in vitro germinability within 15-30 days, while those stored in oils maintained some degree of germinability even after 60 days. Pollen samples stored in oils at -20.degree. C did not show any decline in germinabilty or pollen tube vigour even after 6 months of storage. The results amply demonstrate the feasibility of using oils for short- and long-term pollen storage."
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The following is from my web page:
http://home.neo.rr.com/kuska/pollencollection.htm
Preferably, the dried pollen should be used immediately. If the pollen must be stored, it probably can be kept for several weeks in a refrigerator or for about nine months in a freezer (this is a general statement - how long a given set of pollen will live depends on the variety and even on the growing conditions of the individual plant). After a week or two of refrigerator storage, you may not want to risk using the then questionable pollen on a variety for which you have only a limited number of blooms. Instead, you may want to use the pollen (by itself or mixed with other questionable pollen) on a variety that you have an excess of. Decide on your breeding program ahead of time so that you will be ready to pollinate roses when the first blooms appear in the spring. Try to pollinate when the plants are actively growing and it is cool enough that the plants are not under too much stress. In general, hybridizers do most of their pollinating during the first bloom cycle. The seed on some varieties will not mature when the second bloom cycle is pollinated, especially in northern climates (although 90 days of maturity is considered average, some varieties may take up to 120 days). Unfortunately, some varieties are not as fertile during the first bloom cycle as in later cycles. Also, some varieties mature relatively quickly (as quick as 60 days); and, for these, even the third bloom cycle may be used in warmer areas.
END OF MY WEB PAGE INFORMATION

Here is a link that might be useful: my pollen method