Method for measuring pollen density by improved density gradient centrifugation method

A technology of density gradient centrifugation and concentration gradient, which is applied in the field of plant physiology, can solve the problems of no related reports on the determination of pollen density, and achieve the effects of simple operation, accurate results and good repeatability

Active Publication Date: 2013-01-23
YUNNAN ACAD OF TOBACCO AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[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] The pollen density of tobacco N.alata was measured.

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

Embodiment 2

[0073] The pollen density of tobacco variety N. rustica was measured.

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

Embodiment 3

[0089] The pollen density of tobacco variety N.repanda was measured.

[0090] First, prepare sucrose solutions with mass percentages of 10%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, and place them in a beaker for later use. Secondly, weigh 0.005g of N.repanda pollen and mix it in 500μl of 10% sucrose solution. Once again, first use a syringe to add 50μl of 10% sucrose solution containing pollen samples to the bottom of a 1.5ml centrifuge tube, then suck the high-concentration sucrose solution, insert the syringe into the bottom of the tube, add the high-concentration sucrose solution, and wait for the high-concentration solution to reduce the concentration. From the top of the low solution to the upper layer, add 50μl each of 20%, 30%, 40%, 50%, 55%, 60% and 65% sucrose solution according to the above method to make 10%~65% density gradient centrifugal solution . In a Microfuge 22R centrifuge, centrifuge at 10000 rpm at 20°C for 5 minutes and then let it stand. After the po...

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Abstract

The invention discloses a method for measuring the pollen density by an improved density gradient centrifugation method. The method comprises the following steps of: dissolving cane sugar into water, preparing gradient solutions with different concentrations by mass percentage, mixing pollen to be measured into 10 percent crane sugar solution, and injecting the solution into the bottom of a centrifugation pipe; and then preparing a 20-70% crane sugar density gradient centrifugation solution by a layering method, centrifugating at the speed of 4,000 to 10,000rpm for 5 to 15 min, after the pollen reaches settlement balance, keeping the pollen still, taking crane sugar solution with concentration corresponding to the layer wherein the pollen is suspended, drawing an absorbance standard curve of the pollen to be measured in the crane sugar solution with the corresponding concentration, measuring an absorbance value of the crane sugar solution at the layer an which the pollen is suspended by taking the curve as a reference through a spectrophotometer under a wavelength of 290nm, and calculating the pollen density according to the formula shown in the specification. The method is suitable for measurement of various pollen densities; a result is accurate; the repetitiveness is high; the method is easy to operate and is feasible; and the number of demanded samples is small.

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 Applications(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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