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Method for measuring pollen density by improved density gradient centrifugation method

A technology of density gradient centrifugation and concentration gradient, applied in the field of plant physiology, can solve the problems of no relevant reports on the determination of pollen density, difficulty in collecting pollen, inconvenient operation, etc., and achieve the effect of simple operation, small sample amount and accurate results

Active Publication Date: 2014-09-17
YUNNAN ACAD OF TOBACCO AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Using this method to measure the density of pollen, first of all, it is inconvenient to operate; special instruments are required
Second, this method requires a relatively large amount of pollen, which is difficult and expensive to collect
[0004] The main purpose of density centrifugation is mainly used for the separation of substances, and there is no relevant report on the determination of pollen density

Method used

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  • Method for measuring pollen density by improved density gradient centrifugation method
  • Method for measuring pollen density by improved density gradient centrifugation method
  • Method for measuring pollen density by improved density gradient centrifugation method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Determination of pollen density of tobacco N. alata.

[0058]First, prepare sucrose solutions with mass percentages of 10%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 65%, and 70%, respectively, and place them in beakers for later use. Secondly, weigh 0.001g N.alata pollen and mix it in 100μl 10% sucrose solution. Again, first use a syringe to add 100 μl of 10% sucrose solution containing pollen samples to the bottom of a 1.5ml centrifuge tube, then draw the high-concentration sucrose solution, insert the syringe into the bottom of the tube, add high-concentration sucrose solution, and wait until the concentration of the solution is high. From the top of the lower solution to the upper layer, add 150 μl each of 20%, 30%, 40%, 50%, 55%, 60% and 65% sucrose solution in turn according to the above method to make a density gradient centrifugal solution of 10%~65% . In a Microfuge 22R centrifuge, at 20°C, at a speed of 6000rpm, centrifuge for 10 minutes and then let it stand still...

Embodiment 2

[0073] Determination of pollen density of tobacco variety N. rustica.

[0074] First, prepare sucrose solutions with mass percentages of 10%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 65%, and 70%, respectively, and place them in beakers for later use. Secondly, weigh 0.01g N.rustica pollen and mix it in 1000μl 10% sucrose solution. Again, first add 100 μl of 10% sucrose solution containing pollen samples to the bottom of the 2.0ml centrifuge tube with a syringe, then draw the high-concentration sucrose solution, insert the syringe into the bottom of the tube, add the high-concentration sucrose solution, and wait until the concentration of the solution is high From the top of the lower solution to the upper layer, add 200 μl each of 20%, 30%, 40%, 50%, 55%, 60% and 65% sucrose solution in turn according to the above method to make a 10%~65% density gradient centrifugal solution . In a Microfuge 22R centrifuge, at 20°C, at a speed of 4000rpm, centrifuge for 15 minutes and then let i...

Embodiment 3

[0089] Determination of pollen density of tobacco variety N. repanda.

[0090] First, prepare sucrose solutions with mass percentages of 10%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 65%, and 70%, respectively, and place them in beakers for later use. Secondly, weigh 0.005g N.repanda pollen and mix it in 500μl 10% sucrose solution. Again, first use a syringe to add 50 μl of 10% sucrose solution containing pollen samples to the bottom of the 1.5ml centrifuge tube, then draw the high-concentration sucrose solution, insert the syringe into the bottom of the tube, add high-concentration sucrose solution, and wait until the concentration of the solution is high. From the top of the lower solution to the upper layer, add 50 μl each of 20%, 30%, 40%, 50%, 55%, 60% and 65% sucrose solution in turn according to the above method to make a 10%~65% density gradient centrifugal solution . In a Microfuge 22R centrifuge, at 20°C, at a speed of 10,000rpm, centrifuge for 5 minutes and then let it ...

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Abstract

The invention discloses an improved density gradient centrifugation method for measuring pollen density. Firstly, sucrose is dissolved in water to prepare concentration gradient solutions with different mass percentages, then the pollen to be tested is mixed into 10% sucrose solution, and injected into the bottom of the centrifuge tube. Then, 20-70% sucrose density gradient centrifugation solution was prepared by layer laying method. Centrifuge at 4000~10000rpm for 5~15min at 20°C, wait for the pollen to settle and let it stand still, take the sucrose solution of the corresponding concentration in the layer where the pollen is suspended, and draw the absorbance standard of the pollen to be tested in the sucrose solution of the corresponding concentration Curve, with this curve as a reference, using a spectrophotometer at a wavelength of 290nm, measure the absorbance value of the sucrose solution in the layer where the pollen is suspended, and calculate the pollen density according to the following formula: the present invention is applicable to the measurement of various pollen densities, and the results are accurate and repeatable Good performance, simple operation, easy operation, less sample required.

Description

technical field [0001] The invention belongs to the field of plant physiology, and in particular relates to an improved density gradient centrifugation method for measuring pollen density. Background technique [0002] Pollen is a powdery substance, and the density of pollen is one of the important indicators of pollen physical characteristics. Determining the density of pollen can measure the internal quality of pollen, and the quality of pollen directly affects the quality of plant seeds. Therefore, the determination of pollen physical density can be applied to the detection of the quality of pollen for plant propagation. Determining the density of pollen can also provide a scientifically feasible method for screening pollen. [0003] A conventional method for measuring the density of a powdery substance is to put the powdery substance into a certain amount of Lees bottle containing a liquid medium, and make the liquid medium fully immerse the powdery particles. Accordi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N9/24
Inventor 马文广姚恒牛永志郑昀晔索文龙宋碧清李元君
Owner YUNNAN ACAD OF TOBACCO AGRI SCI
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