CN-121970681-A - Tissue culture method for reducing cercis chinensis browning

Abstract

The invention discloses a tissue culture method for reducing cercis chinensis browning, which comprises the steps of S1, cleaning and sterilizing a cercis chinensis stem section with axillary buds, taking the axillary buds in a sodium sulfite solution, S2, performing primary culture on the axillary buds to obtain aseptic seedlings, and performing anti-browning culture and growth promotion culture on the aseptic seedlings to obtain the cercis chinensis tissue culture seedlings, wherein culture medium components used in the anti-browning culture comprise improved MS, IBA, 2,4-D, rutin and sodium sulfite. Compared with the prior art, the invention improves the sterile seedling obtaining rate by 81.67 percent through improving the disinfection method, obtains the cercis chinensis sterile seedling transplanting survival rate of 88.33 percent through anti-browning growth promotion culture, and the surviving cercis chinensis sterile seedling leaves are dark green or cyan, and the stems are more robust.

Inventors

• ZHANG XIAONING
• LIN DONG
• LU JUAN
• SUN XI
• ZHANG YE
• LIU HAILONG
• QIN ZIHAI
• LI LILI
• LU XUYA
• LI JIANFAN

Assignees

• 广西壮族自治区林业科学研究院
• 玉林市林业科学研究所

Dates

Publication Date

20260505

Application Date

20251218

Priority Date

20241219

Claims (7)

1. 1. A tissue culture method for reducing cercis chinensis browning, the tissue culture method comprising: s1, cleaning and disinfecting a stem section of cercis chinensis with axillary buds, and taking the axillary buds in a sodium sulfite solution, wherein the specific steps are as follows: s11, taking a cercis chinensis branch containing 2-3cm axillary buds; S12, soaking the redbud branches in alcohol for 30-120S, then washing with distilled water, and soaking in H 2 O 2 for 5-8 min, then washing with distilled water; S13, taking off 2-3cm axillary buds on a cercis chinensis branch in a sodium sulfite solution to obtain the axillary buds, wherein the sodium sulfite solution is obtained by adding 5-7 mu L of 1mg/L sodium sulfite into 20-40mL of distilled water; S2, performing primary culture on the axillary buds to obtain aseptic seedlings, performing anti-browning growth promotion culture on the aseptic seedlings, taking stem segments of the aseptic seedlings, performing secondary culture on the stem segments of the aseptic seedlings, wherein a culture medium component used in the anti-browning culture consists of modified MS+1-2mg/L IBA+2-4 mg/L2, 4-D+1-2mg/L L-cysteine+1-2 mg/L brassinosteroid+1-2 mg/L sodium sulfite, and a culture medium component used in the primary culture consists of WPM+1-2mg/L sodium sulfite.
2. 2. A tissue culture method for reducing cercis chinensis browning according to claim 1, wherein the modified MS medium comprises the specific components of 825mg/L ammonium nitrate, 950mg/L potassium nitrate, 220mg/L calcium chloride, 185mg/L magnesium sulfate, 85mg/L potassium dihydrogen phosphate, 22.3mg/L manganese sulfate, 8.6mg/L zinc sulfate, 6.2mg/L boric acid, 0.83mg/L potassium iodide, 0.25mg/L sodium molybdate, 0.025mg/L copper sulfate, 0.025mg/L cobalt chloride, 37.3mg/L disodium ethylenediamine tetraacetate, 27.8mg/L ferrous sulfate, 100mg/L inositol, 2.0mg/L glycine, 0.1mg/L thiamine hydrochloride, 0.5mg/L pyridoxine hydrochloride, and 0.5mg/L niacin.
3. 3. The tissue culture method for reducing cercis chinensis browning according to claim 1, wherein the culture conditions of the anti-browning growth-promoting culture are that the culture is carried out after the culture is carried out under the conditions that the culture temperature is 25+/-2 ℃ and the illumination intensity is 3000-4000Lx and the illumination time is 12-14 h/d until the height of the sterile seedlings is 5-7 cm.
4. 4. The tissue culture method for reducing browning of cercis chinensis according to claim 1, wherein the length of the branches of cercis chinensis is 3-5cm between adjacent internodes of the stem node where the 2-3cm axillary bud is located.
5. 5. The tissue culture method for reducing cercis browning according to claim 1, wherein said H 2 O 2 concentration is 10-12%.
6. 6. The tissue culture method for reducing browning of cercis chinensis according to claim 1, wherein the culture conditions of the initial culture are that the culture temperature is 18+/-2 ℃ and the light intensity is 500-1000Lx, and the light time is 6-8 h/d for 4-5 days.
7. 7. The tissue culture method for reducing browning of cercis chinensis according to claim 1, wherein in the step of subjecting the aseptic seedlings to anti-browning growth-promoting culture, the aseptic seedlings are transplanted as a whole.

Description

Tissue culture method for reducing cercis chinensis browning Technical Field The invention relates to the technical field of plant tissue culture, in particular to a tissue culture method for reducing cercis chinensis browning. Background Although natural protection areas are established for protecting the cercis chinensis for a long time, the wild resources of the cercis chinensis still have a trend of drastically reducing, and plant tissue culture is necessary for preserving germplasm resources of the cercis chinensis. To achieve protection of the germplasm resources of cercis chinensis, the present inventors have focused on obtaining cercis chinensis tissue culture seedlings by plant tissue culture. However, during the tissue culture of cercis chinensis, the inventors found that the tissue culture of cercis chinensis had the following difficulties: (1) Serious pollution and browning of the axillary buds of the redbud tree occur in the primary culture stage, so that the aseptic seedlings are low in yield; (2) The inoculated axillary buds or stem segments of the redbud tree are dyed into light yellow or brown by taking the inoculating points as dots in the culture process, and the inoculating materials are also brown to different degrees, and the growth is affected by light brown, so that the inoculated materials die seriously due to withering; (3) The slow growth of the stem node of the stem section of the cercis chinensis leads to long culture time and difficulty in sprouting larger leaves. The information disclosed in this background section is only for enhancement of understanding of the general background of the invention and should not be taken as an acknowledgement or any form of suggestion that this information forms the prior art already known to a person of ordinary skill in the art. Disclosure of Invention The invention mainly aims to provide a tissue culture method for reducing cercis chinensis test tube plantlet browning, which aims to solve the problem that wild resources of cercis chinensis are gradually reduced and the resources of cercis chinensis germplasm are possibly lost, and overcome the difficulties in the cercis chinensis plant tissue culture process in the background technology. In order to achieve the above object, the present invention provides a tissue culture method for reducing cercis chinensis browning, the tissue culture method comprising: S1, cleaning and disinfecting stem sections of the cercis chinensis with axillary buds, and taking the axillary buds in a sodium sulfite solution; S2, performing primary culture on the axillary buds to obtain aseptic seedlings, performing anti-browning growth promotion culture on the aseptic seedlings, and taking stem segments of the aseptic seedlings for secondary culture, wherein culture medium used in the anti-browning culture comprises improved MS, IBA, 2,4-D, L-cysteine, rutin and sodium sulfite. Preferably, in the technical scheme, the culture medium used for the anti-browning culture consists of modified MS+1-2mg/L IBA+2-4 mg/L2, 4-D+1-2mg/L L-cysteine+1-2 mg/L brassinosteroid+1-2 mg/L sodium sulfite. Preferably, in the technical scheme, the culture conditions of the anti-browning growth-promoting culture are that the culture is carried out after the culture is carried out to the height of 5-7cm under the conditions that the culture temperature is 25+/-2 ℃ and the illumination intensity is 3000-4000Lx and the illumination time is 12-14 h/d. Preferably, in the above technical solution, in step S1, the step of cleaning and disinfecting the stem segments of cercis chinensis with axillary buds, and the specific steps of taking the axillary buds in sodium sulfite solution are as follows: s11, taking a cercis chinensis branch containing 2-3cm axillary buds; S12, soaking the redbud branches in alcohol for 30-120S, then washing with distilled water, and soaking in H 2O2 for 5-8 min, then washing with distilled water; S13, taking off 2-3cm axillary buds on the redbud branches in sodium sulfite solution to obtain the axillary buds. Preferably, in the above technical scheme, the length of the redbud branches is 3-5cm between adjacent nodes of the stem node where the 2-3cm axillary bud is reserved. According to the technical scheme, the method protects the axillary buds by reserving the 3-5cm between adjacent internodes of the stem node where the axillary buds of 2-3cm are positioned, and avoids the damage of cell membranes of cells near the incision caused by disinfectant in the disinfection process of the cercis chinensis explant, so that the death of the explant caused by the damage of the axillary buds is reduced. Preferably, in the above technical solution, the concentration of H 2O2 is 10-12%. Preferably, in the above technical scheme, the sodium sulfite solution is obtained by adding 5-7 mu L of 1mg/L sodium sulfite into 20-40mL of distilled water. Preferably, in the above technical scheme, the culture medium used for the primary culture