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Sodium azide mutagenizing method for juvenile citrus internodal stem segments

A technology of sodium azide, treatment method, applied in horticultural methods, botanical equipment and methods, chemicals for biological control, etc., can solve problems such as difficult mutants, instability, etc. The effect of simplicity and easy detection method

Active Publication Date: 2018-07-24
HUNAN INST OF GARDENING
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because citrus fruit trees have a long childhood period (generally more than 5 years), citrus seeds are treated with sodium azide (causing genetic material variation in germ cells, and then forming stable inheritance through sexual reproduction) to obtain stable mutations from their offspring The working cycle of the body needs a long time; in addition, the axillary buds of citrus are already in a certain state of differentiation, and using sodium azide to directly treat the axillary buds (scions) of citrus often only causes mutations in a few cells in each meristematic cell of the axillary buds. This type of mutation has the characteristics of instability, that is, the new shoot leaves that germinate in the leaf axils of many mutant leaves return to the normal state, so it is also difficult to obtain mutants with stable mutant traits

Method used

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  • Sodium azide mutagenizing method for juvenile citrus internodal stem segments
  • Sodium azide mutagenizing method for juvenile citrus internodal stem segments
  • Sodium azide mutagenizing method for juvenile citrus internodal stem segments

Examples

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Embodiment 1

[0045] A sodium azide mutagenesis treatment method for juvenile citrus internode stem segments, comprising the steps of:

[0046] Step 1. Mutagenesis treatment solution configuration: Weigh an appropriate amount of sodium azide and add it to a 0.1mol / L potassium phosphate buffer solution with a pH value of 3.00 (±0.05), and configure the final concentrations to be 0mmol / L and 0.5mmol / L respectively. Mmol / L, 1mmol / L, 2mmol / L and 5mmol / L mutagenesis treatment solutions. When using mutagenesis treatment solution, add surfactant Silwet L-77 (final concentration: 0.5%).

[0047] Step 2. Peel off the outer testa of the citrus seeds in the ultra-clean workbench, sterilize with 1% NaClO for 10 minutes, discard the NaClO liquid in the ultra-clean workbench, rinse 3 times with sterile water and place them overnight at 4°C before sowing in MS medium (pH 5.7). After sowing, cultivate in the dark for about 20 days, and then culture in the light. The light culture conditions are: 12h ligh...

Embodiment 2

[0055] A sodium azide mutagenesis treatment method for juvenile citrus internode stem segments, comprising:

[0056] Step 1. Mutagenesis treatment solution configuration: Weigh an appropriate amount of sodium azide and add it to a 0.1mol / L potassium phosphate buffer solution with a pH value of 3.00 (±0.05) to configure a mutagenesis solution with a final concentration of 2mmol / L treatment fluid. When using mutagenesis treatment solution, add surfactant Silwet L-77 (final concentration: 0.5%).

[0057] Step 2. Peel off the outer testa of the citrus seeds in the ultra-clean workbench, sterilize with 1% NaClO for 10 minutes, discard the NaClO liquid in the ultra-clean workbench, rinse 3 times with sterile water and place them overnight at 4°C before sowing in MS medium (pH 5.7). After sowing, cultivate in the dark for about 20 days, and then culture in the light. The light culture conditions are: 12h light / 12h dark, light intensity 40μmol·m - 2 the s -1 , cultured for 10 day...

Embodiment 3

[0062] A sodium azide mutagenesis treatment method for juvenile citrus internode stem segments, comprising:

[0063] Step 1. Mutagenesis treatment solution configuration: Weigh an appropriate amount of sodium azide and add it to a 0.1mol / L potassium phosphate buffer solution with a pH value of 3.00 (±0.05) to configure a mutagenesis solution with a final concentration of 1mmol / L treatment fluid. When using mutagenesis treatment solution, add surfactant Silwet L-77 (final concentration: 0.5%).

[0064] Step 2. Peel off the outer testa of the citrus seeds in the ultra-clean workbench, sterilize with 1% NaClO for 10 minutes, discard the NaClO liquid in the ultra-clean workbench, rinse 3 times with sterile water and place them overnight at 4°C before sowing in MS medium (pH 5.7). After sowing, cultivate in the dark for about 20 days, and then culture in the light. The light culture conditions are: 12h light / 12h dark, light intensity 40μmol·m - 2 the s -1 ) 10d, the growth tem...

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Abstract

The invention discloses a sodium azide mutagenizing method for juvenile citrus internodal stem segments, comprising: I, taking citrus young plantlets more than 1 mm in stem thickness and 5-10 cm in plantlet height, cutting to obtain internodal stem segments, soaking the internodal stem segments in a mutagenizing liquid, mutagenizing in darkness for 1-3 h, wherein the mutagenizing liquid includes sodium azide having a concentration of 0.5-2 mmol / L; II, subjecting the mutagenized internodal stem segments to tissue culture until sectional wounds of the internodal stem segments grow regenerated buds; III, using an SRAP (sequence-related amplified polymorphism) molecular markup method to detect the regenerated buds that vary, and subjecting the varied regenerated buds to rooting culture to obtain complete citrus variant plants. It is clarified that sodium azide-mutagenized juvenile citrus internodal stem segments may produce stable variant plants. The method of the invention has mutagenizing efficiency of greater than 10% and has significant mutagenizing effect and good operating convenience.

Description

technical field [0001] The invention belongs to the technical field of creating citrus variation, in particular to a sodium azide mutagenesis treatment method for juvenile citrus internode stem segments. Background technique [0002] Sodium azide is a non-toxic, relatively safe chemical mutagen. After the branches of Newhall navel orange were treated with sodium azide, there were leaves with morphological variation on the new shoots from some leaf axils of the branches, and molecular markers could be used to detect the difference in genome composition between the mutant leaves and the control leaves, These results indicate that sodium azide has a mutagenic effect on the genetic material of citrus cells. Because citrus fruit trees have a long childhood period (generally more than 5 years), citrus seeds are treated with sodium azide (causing genetic material variation in germ cells, and then forming stable inheritance through sexual reproduction) to obtain stable mutations fr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01G17/00A01H4/00A01N65/40A01N43/38A01N37/10A01N59/00A01P21/00
CPCA01G17/005A01H4/00A01H4/008A01N37/10A01N43/38A01N59/00A01N65/00A01N65/12A01N65/40
Inventor 孔佑涵吴娟娟李先信易春苑平张文谢新华杨辉学
Owner HUNAN INST OF GARDENING
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