Manufacture method of continuous paraffin sections for gonad of pelodiscus sinensis in embryonic period and application of manufacture method in sex determination

A production method and paraffin section technology are applied in the preparation of test samples, measuring devices, and material analysis by optical means, etc., which can solve problems such as difficulty in determining the gender of Chinese soft-shelled turtles, and achieve easy popularization and application, release of space resistance, and difficulty in The effect of missing samples

Inactive Publication Date: 2014-07-23
ZHEJIANG WANLI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] From the above analysis, it can be seen that there are great difficulties in judging the sex of soft-shelled turtles by anatomical methods without long-term breeding.

Method used

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  • Manufacture method of continuous paraffin sections for gonad of pelodiscus sinensis in embryonic period and application of manufacture method in sex determination
  • Manufacture method of continuous paraffin sections for gonad of pelodiscus sinensis in embryonic period and application of manufacture method in sex determination
  • Manufacture method of continuous paraffin sections for gonad of pelodiscus sinensis in embryonic period and application of manufacture method in sex determination

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041]①Sample fixation: After 24 days of incubation, collect 100 soft-shelled turtle embryos with an incubation temperature of 33°C. After cutting the abdominal cavity, take out the gonad-mesonephric-adrenal complex. It should be noted that only one side of the gonads on both sides of the soft-shelled turtle needs to be taken out. That is, the taken-out complex was immediately put into 4% paraformaldehyde fixative solution for fixation, and then transferred to 70% alcohol solution for storage after fixing for 3 hours;

[0042] ② Gonad staining: the gonads collected in step ① were first stained with eosin, and the eosin staining time was 2 minutes, and rinsed with 80% alcohol after eosin staining;

[0043] ③Dehydration and transparency procedure: Dehydrate the soft-shelled turtle gonads in step ② with gradient ethanol; soak in 80% ethanol for 20 minutes, 90% ethanol for 20 minutes, and 100% ethanol for 10 minutes; then use absolute ethanol with a volume ratio of 1:1 : Transitio...

Embodiment 2

[0051] ①Sample fixation: After 31 days of incubation, collect 100 Chinese soft-shelled turtle embryos with an incubation temperature of 32°C. After cutting the abdominal cavity, take out the gonad-mesonephric-adrenal complex. It should be noted that only one side of the soft-shelled turtle needs to be taken out. That is, the taken-out complex was immediately put into formalin fixative for fixation, and after fixation for 24 hours, it was transferred to 80% alcohol solution for storage;

[0052] ② Gonad staining: the gonads collected in step ① were first stained with eosin, and the eosin staining time was 2 minutes, and rinsed with 70% alcohol after eosin staining;

[0053] ③Dehydration and transparency procedure: Dehydrate the soft-shelled turtle gonads in step ② with graded ethanol; soak in 80% ethanol for 40 minutes, 90% ethanol for 30 minutes, and 100% ethanol for 20 minutes; then use absolute ethanol with a volume ratio of 1:1 : Transition treatment with xylene for 20 minu...

Embodiment 3

[0060] ①Sample fixation: Collect 100 Chinese soft-shelled turtle embryos with an incubation temperature of 31°C 1 day after hatching, cut the abdominal cavity and take out the gonad-mesonephric-adrenal complex. Immediately put the taken-out complex into Born's fixative solution, fix it for 12 hours, and then transfer it to 75% alcohol solution for storage;

[0061] ② Gonad staining: the gonads collected in step ① were first stained with eosin, and the eosin staining time was 3 minutes, and rinsed with 90% alcohol after eosin staining;

[0062] ③Dehydration and transparency procedure: Dehydrate the soft-shelled turtle gonads in step ② with gradient ethanol; soak in 80% ethanol for 50 minutes, 90% ethanol for 15 minutes, and 100% ethanol for 30 minutes; then use absolute ethanol with a volume ratio of 1:1 : Transition treatment with xylene for 30 minutes, then soak in xylene for 30 minutes to make it transparent;

[0063] ④ Paraffin penetration and embedding: Transition treatme...

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Abstract

The invention relates to a manufacture method of continuous paraffin sections for gonad of pelodiscus sinensis in embryonic period and application of the manufacture method in sex determination. The manufacture method comprises the following steps: (1) fixing a sample, (2) dying the gonad, (3) carrying out dehydration and transparentizing procedure, (4) absorbing paraffin and embedding, (5) cutting into sections and flatting sections, and (6) dying and mounting. The manufacture method is characterized in that the gonad is first dyed in the step (2); sections are subjected twice flatting by adopting water bath and high-temperature paraffin in the step (5). Compared with the prior art, the manufacture method has the advantages that the gonad is dyed first before being embedded, the gonads of a plurality of individuals are embedded in one paraffin block, determination is carried out through a manner of continuous tissue sections, and then the result accuracy is ensured. The manufacture method is simple to operate and convenient to popularize and apply. By utilizing the manufacture method, sex ratio of the pelodiscus sinensis in the embryonic period can be rapidly and accurately subjected to statistics.

Description

technical field [0001] The invention relates to a method for making continuous paraffin slices and its application, in particular to a method for making paraffin slices of gonads at the embryonic stage of soft-shelled turtle and its application in judging the sex of embryos. Background technique [0002] Chinese soft-shelled turtle (Pelodiscus sinensis), also known as soft-shelled turtle, bastard, water fish, yuan fish, turtle fish, foot fish and old turtle, belongs to the subphylum Vertebtata, class Reptilia, order Chelonia, Trionychidae (Trionychidae), widely distributed in my country except Ningxia, Xinjiang, Qinghai and Tibet, is more common in the Yangtze River Basin and South China. It is the most widely cultivated aquatic reptile in my country and has important economic and medicinal value. Value; abroad distributed in Japan, Korea and Vietnam. Regarding the embryonic development of reptiles, different hatching conditions and hormone treatments can affect the gonad dif...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/28G01N21/84
Inventor 王伟钱国英李彩燕宋伟汪财生尹尚军朴龙斗
Owner ZHEJIANG WANLI UNIV
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