CN115251007A - New strain naked rat, and cultivation method and application thereof - Google Patents
New strain naked rat, and cultivation method and application thereof Download PDFInfo
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- 238000012364 cultivation method Methods 0.000 title abstract description 4
- 238000011160 research Methods 0.000 claims abstract description 19
- 238000009396 hybridization Methods 0.000 claims abstract description 9
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- 238000009395 breeding Methods 0.000 claims description 17
- 230000001488 breeding effect Effects 0.000 claims description 10
- 238000000034 method Methods 0.000 claims description 8
- 206010061598 Immunodeficiency Diseases 0.000 claims description 2
- 208000029462 Immunodeficiency disease Diseases 0.000 claims description 2
- 230000007813 immunodeficiency Effects 0.000 claims description 2
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- 210000003128 head Anatomy 0.000 abstract description 9
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- 108090000623 proteins and genes Proteins 0.000 description 13
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- 238000013112 stability test Methods 0.000 description 2
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
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Abstract
The invention relates to a new strain naked rat, a cultivation method and application thereof. The new strain naked rat is produced by taking an NIH naked rat as an original male parent and an SD rat as an original female parent through successive generations of close-up hybridization, eyes of the new strain naked rat are red, the skin of the whole body of the new strain naked rat is white, white hairs on the head, the back and the tail of the new strain naked rat periodically fall off in a growing period, the new strain naked rat is different from various naked rat strains used in the current research field, the thymus volume of the new strain naked rat is smaller than that of the NIH naked rat through continuous observation, and the growth speed, the environmental tolerance and the service life length of the new strain naked rat are superior to those of the NIH naked rat.
Description
Technical Field
The invention belongs to the field of breeding of new strain naked rats, and particularly relates to a new strain naked rat, a breeding method and application thereof.
Background
Athymic nude mice (nude mice for short) have almost no immunity in vivo due to congenital thymic deficiency or low function, and have become indispensable experimental animal models in the field of medical research, and particularly have extremely special values in the aspects of stem cell and tissue engineering, oncology, immunology, safety evaluation of medicines and biological products and screening of effective medicines.
According to different scientific research purposes and types, nude mice are divided into nude mice and nude rats, wherein the nude rats have incomparable advantages in the scientific research field due to the characteristics of longer service life, larger body size, stronger tolerance, wider skin area and the like.
Research shows that the service life of a nude rat is longer than that of the nude rat, the nude rat can be transplanted with a tumor, the proliferation characteristics of the tumor can be observed for a longer time, and the nude rat has large body size and sufficient blood volume, so that more blood volume can be collected in the aspect of blood biochemical analysis for scientific research, large-scale operations which cannot be tolerated by some nude rats can be well implemented on the nude rat, in addition, the feeding difficulty of the nude rat is lower than that of the nude rat, the burden of a large number of animals for scientific research personnel can be reduced, and the characteristics in all aspects prove that the nude rat has a very large application prospect in the field of medical research.
Among the numerous nude rats, NIH nude rats are more commonly used. However, the NIH nude rats have relatively short life span and the skin of the whole body is in a state of alternating black and white, so that the scientific research is greatly challenged because the life span is not long enough, the growth speed is slow and the direction of blood vessels is not well distinguished by the black part of the skin color when the continuous long-stage observation research and the skin blood vessel injection are carried out.
Therefore, the technical scheme of the invention is provided.
Disclosure of Invention
In order to solve the problems in the prior art, the invention provides a new strain of nude rats, a breeding method and application thereof. According to the method for breeding the new strain of the naked rat, the NIH naked rat and the SD rat are hybridized to generate offspring, and then continuous close hybridization screening is carried out, so that the new strain of the naked rat which has the advantages of higher growth speed, longer service life, stronger environmental tolerance, smaller thymus and different biological properties (eyes are red, the skin of the whole body is white, white hairs on the head, the back and the tail in the growth period periodically appear and fall off, and no thymus exists) from the existing various naked rat strains is finally obtained.
The scheme of the invention provides a breeding method of a new strain of naked rats, which comprises the following steps:
(1) Selecting male NIH naked rats and female SD rats as primary male parents and primary female parents respectively, and hybridizing in a cage until the F1 generation is born; wherein, F1 generation is about 25d, and 1 week later observation shows that all cubs are heterozygote rats containing SD rat gene and NIH naked rat gene (rats with black eyes, black and white hair and thymus) all over the whole body;
(2) After the F1 generation is sexually mature, carrying out cage crossing on the male and the female of the F1 generation to the F2 generation; wherein, F2 is born for about 25 days, and naked rats with the same appearance characteristics as NIH naked rats (black eyes, black and white hair between the head, the back and the tail periodically appears along with growth and then falls off and no thymus exists) are observed after 1 week, but the gene group of the naked rat has mixed SD rat genes;
(3) After the F2 generation is mature, carrying out cage-combination hybridization on male naked rats with the same appearance characteristics as NIH naked rats in the F2 generation and female rats in the F1 generation to obtain F3 generation in which the new strain naked rats exist. Among these, the new strain of nude rats, which is low in percentage, appeared in the F3 generation (red eyes, white hair on the head, back, and tail, which periodically appeared with growth and then fell off, and thymus-free, existed).
As such, in order to increase the proportion of nude rats in the new strain, the following preferred protocol is followed:
(4) After the F3 generation is sexually mature, selecting the male naked rat in the F3 generation and the female rat in the F1 generation to carry out cage-combination hybridization until the F4 generation is generated; wherein, F4 generation is about 25d, and 1 week later observation shows that rats with the same appearance characteristics as SD rats (rats with red eyes, white hair distributed on the whole body and thymus) can appear in the F4 generation, but the genome of the rats is mixed with NIH naked rat genes;
(5) After the F4 generation is mature, selecting male nude rats in the F3 generation and female rats with the same appearance characteristics as SD rats in the F4 generation for cage crossing, and obtaining the new strain nude rats in the F5 generation.
The inventor repeatedly verifies that more than half of the F5 generation born after about 25d is the new strain naked rat, and the rest is the rat with red eyes, white hair distributed on the whole body and thymus. And then, the genetic stability test is carried out according to the method for breeding F5, and the new strain nude rats can stably appear according to the proportion of more than 1/2.
Preferably, for stability of the incubation, the incubation is performed in an SPF-grade feeding environment at room temperature of 20-26 ℃.
Based on the same technical concept, the invention also provides a new strain of nude rats obtained by the breeding method.
For the purpose of facilitating understanding of the present invention, the technical idea of the present invention will be explained.
In the process of research and research, the biological characteristics of SD rats and NIH naked rats provide important hints for the inventor: NIH naked rats have relatively short service life, the skin of the whole body is in a state of alternating black and white, and the service life is short, the growth speed is slow, and the black part of the skin color is not good for distinguishing the trend of blood vessels when the NIH naked rats are subjected to continuous long-stage observation research and skin blood vessel injection drugs, so that great challenges are brought to scientific research; however, contrary to the above, the SD rat has a long life, strong environmental tolerance, and white skin color, and can be used for long-term scientific observation and research, and can also be used for conveniently injecting various drugs into skin blood vessels, but the SD rat has normal immunity, and does not have the characteristic of no immune rejection in NIH nude rat tumor transplantation. Based on the biological characteristics of the two rats, the inventor hybridizes the SD rat with the NIH naked rat, and finally cultivates the new strain of the NIH naked rat which has the characteristics of no thymus and no immunity of the NIH naked rat, and has the characteristics of high growth speed, long service life, strong environmental tolerance, white whole skin and the like of the SD rat through a continuous close hybridization cultivation mode.
Therefore, the new strain of the nude rat has important significance for animal model research of the nude rat in the medical research field, can enrich the variety types of the nude rat, and can provide more selection ways for establishing an immunodeficiency model.
The invention has the beneficial effects that:
compared with NIH (human immunodeficiency virus) nude rats, the new strain nude rats obtained by the cultivation method have the advantages of higher growth speed, higher tolerance capability and longer life, red eyes, white skin of the whole body, and periodic growth and falling of white hairs at the neck and the tail in the growth period, are different from the existing strain nude rats in the field of scientific research, and have extremely special values in the aspects of safety evaluation of stem cells and tissue engineering, oncology, immunology, medicine and biological products and screening of effective medicines.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the embodiments or the prior art descriptions will be briefly described below, it is obvious that the drawings in the following description are only some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a graph showing the comparison of biological characteristics of SD rats, NIH nude rats, and new strain nude rats.
FIG. 2 is a graph showing the comparison of growth and development rates of SD rats, NIH nude rats, and new strain nude rats (age in week versus length).
FIG. 3 is a graph showing the comparison of growth and development rates (week-old versus body weight) of SD rats, NIH nude rats, and New strain nude rats.
FIG. 4 is a thymus anatomy diagram of SD rats, NIH nude rats, new strain nude rats.
FIG. 5 is an enlarged view of the thymus anatomy of SD rat, NIH nude rat, and new strain nude rat.
FIG. 6 is a graph showing the comparison of the number of female offspring of heterozygote of NIH nude rat and new strain nude rat.
FIG. 7 is a graph showing a comparison of the proportion of heterozygote female nude mice appearing in NIH nude rats and new strain nude rats.
FIG. 8 is a comparison of the length of life of NIH nude rats, new strain nude rats, in a common feeding environment (non-SPF class, environment with potential contaminants and conditionally pathogenic pathogens).
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention will be described in detail below. It should be apparent that the described embodiments are only some embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the examples given herein without any inventive step, are within the scope of the present invention.
Example 1
This example provides a method for breeding new strain of nude rats in an SPF-class breeding environment at a constant temperature of 20 ℃, comprising the steps of:
(1) Selecting male NIH naked rats and female SD rats as a primary male parent and a primary female parent respectively, and hybridizing in a cage until the F1 generation is born; wherein, the F1 generation is about 25d, 1 week later observation shows that all cubs are heterozygote rats containing SD rat gene and NIH naked rat gene (rats with black eyes, black and white hair and thymus) all around;
(2) After the F1 generation is sexually mature, carrying out cage crossing on males and females of the F1 generation to the F2 generation; wherein, F2 is born for about 25d, and after 1 week, naked rats with appearance characteristics similar to NIH naked rats (black eyes, black and white hair on the head, back and tail part periodically appear along with growth and then fall off, and thymus-free exists) appear at probability, but the genome of the naked rats is mixed with SD rat genes;
(3) After the F2 generation is mature, carrying out cage-combination hybridization on male naked rats with the same appearance characteristics as NIH naked rats in the F2 generation and female rats in the F1 generation to obtain F3 generation in which the new strain naked rats exist. Among these, the new strain of nude rats, which is low in percentage, appeared in the F3 generation (red eyes, white hair on the head, back, and tail, which periodically appeared with growth and then fell off, and thymus-free, existed).
Example 2
This example provides a preferred breeding method for new strains of nude rats in an SPF-class breeding environment thermostated at 26 ℃, comprising the following steps:
(1) Selecting male NIH naked rats and female SD rats as primary male parents and primary female parents respectively, and hybridizing in a cage until the F1 generation is born; wherein, F1 generation is about 25d, and 1 week later observation shows that all cubs are heterozygote rats containing SD rat gene and NIH naked rat gene (rats with black eyes, black and white hair and thymus) all over the whole body;
(2) After the F1 generation is sexually mature, carrying out cage crossing on males and females of the F1 generation to the F2 generation; wherein, F2 is born for about 25 days, and naked rats with the same appearance characteristics as NIH naked rats (black eyes, black and white hair between the head, the back and the tail periodically appears along with growth and then falls off and no thymus exists) are observed after 1 week, but the gene group of the naked rat has mixed SD rat genes;
(3) After the F2 generation is mature, carrying out cage-combination hybridization on male naked rats with the same appearance characteristics as NIH naked rats in the F2 generation and female rats in the F1 generation to obtain F3 generation in which the new strain naked rats exist. Among these, the new strain of nude rats, which is low in percentage, appeared in the F3 generation (red eyes, white hair on the head, back, and tail, which periodically appeared with growth and then fell off, and thymus-free, existed).
(4) After the F3 generation is mature, selecting the male naked rat and the F1 generation female rat in the F3 generation to perform cage-combining hybridization until the F4 generation is born; wherein, the F4 generation is about 25 days, and 1 week later observation shows that rats with the same appearance characteristics as SD rats (rats with red eyes, white hair distributed on the whole body and thymus) can appear in the F4 generation, but the genome of the rats is mixed with NIH nude rat genes;
(5) After the F4 generation is sexually mature, selecting the male nude rat in the F3 generation and the female rat with the same external features as the SD rat in the F4 generation to perform cage crossing, and obtaining the new strain nude rat in the F5 generation.
It should be noted that, through repeated verification by the inventor, more than half of the F5 generation born after about 25d is the new strain of naked rats, and the rest are rats with red eyes, white hair distributed on the whole body and thymus. And then, the genetic stability test is carried out according to the method for breeding F5, and the new strain nude rats can stably appear according to the proportion of more than 1/2.
Test examples
To verify the various indicators of the new strain of nude rats according to the present invention, the following comparative experiments were performed.
Comparison 1: and (5) comparing biological characteristics.
Images were taken of post-mortem SD rats, NIH nude rats, and new strain nude rats described in the present invention, as shown in FIG. 1.
As is evident from FIG. 1, SD rats have the largest body type and the skin color is white; NIH naked rat has minimal body type, and head skin and hair are obviously black; the new strain of nude rats has a centered body type, and the whole skin is white, showing the same properties as those of SD rats.
Comparison 2: growth and development speed contrast chart-relationship between week age and body length.
Statistics were made on the relationship between the week age and length of SD rats, NIH nude rats and new strain nude rats described in the present invention, as shown in fig. 2.
As is evident from fig. 2, the SD rats consistently have the longest length and the NIH nude rats consistently have the shortest length over time, while the new strain of nude rats has the middle length, which is consistent with the information reflected in the photograph of fig. 1.
Comparison 3: growth and development speed contrast chart-relationship between week age and body weight.
Statistics were made on the relationship between the week age and body weight of SD rats, NIH nude rats and new strain nude rats described in the present invention, as shown in fig. 3.
As is evident from FIG. 3, the weight of SD rats was consistently highest and the weight of NIH nude rats was consistently lowest over time, with the new strain of nude rats being in the middle. This is consistent with the common sense conclusion of "weight and length being positively correlated", in concert with figure 2.
The information in fig. 2 and fig. 3 shows that the growth and development speed of the new strain of nude rats is significantly faster than that of the existing NIH nude rats.
Comparison 4: thymus size comparison.
SD rats, NIH nude rats and the new strain nude rats of the present invention were dissected and directly observed for thymus size, as shown in FIGS. 4 and 5.
As is apparent from FIGS. 4 and 5, the thymus of SD rats is normal, the thymus of NIH nude rats is atrophied, and the thymus of the new strain nude rats is further atrophied. The new strain of nude rats is shown to have athymic and nonimmunity characteristics in characteristic. And the thymus is smaller than that of NIH nude rats, which indicates that the nude rats have better nude mouse characteristics.
Comparison No. 5: and comparing the reproductive capacity.
As can be seen from FIG. 6, in general, female nude rats are used as female parents for reproduction because of their extremely weak reproductive ability, and no statistically significant difference was found by comparing the number of single-birth born between the female parent of the new line nude rat heterozygote of the present invention and the female parent of NIH nude rat heterozygote.
With reference to fig. 7, the proportion of nude mice appearing in the new strain nude rat heterozygote female parent single-fetus pup is found to be higher than that of the NIH nude rat heterozygote female parent through observation, and the finding can prove that the new strain nude rat can produce more nude rats in the same time in the breeding process, so that the time cost for breeding the mice is greatly saved for researchers.
Comparison 6: and comparing the life lengths (the survival rate is counted according to months, and the survival conditions are the same).
As is evident from fig. 8, when fed in a normal environment (non-SPF grade, potential pollutants and conditionally pathogenic pathogens present in the environment), the survival rate for NIH nude rats was statistically reduced to about 90% in dead individuals at 3 months of feeding, and the survival rate gradient was reduced during 3 months to 6 months, with a 4 month survival rate of about 60%, a 5 month survival rate of about 40%, and a 6 month survival rate of about 30%; in a contrary view of the new strain of nude rats, which can be intuitively obtained from fig. 8, the survival rate curve is always higher than that of NIH nude rats, dead individuals appear only when the rats are counted at the 5 th month, the survival rate is about 90 percent at the moment, the survival rate is reduced to about 70 percent at the 6 th month, and the survival rate is still obviously higher than that of the NIH nude rats by about 30 percent. Therefore, the new strain of the naked rat has longer survival capability and stronger tolerance.
The above description is only for the specific embodiments of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art can easily conceive of the changes or substitutions within the technical scope of the present invention, and all the changes or substitutions should be covered within the scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the appended claims.
Claims (6)
1. A method for breeding a new strain of nude rats is characterized by comprising the following steps:
(1) Selecting male NIH naked rats and female SD rats as a primary male parent and a primary female parent respectively, and hybridizing in a cage until the F1 generation is born;
(2) After the F1 generation is sexually mature, carrying out cage crossing on males and females of the F1 generation to the F2 generation;
(3) After the F2 generation is sexually mature, male nude rats with the same appearance characteristics as NIH nude rats in the F2 generation and female rats in the F1 generation are subjected to cage crossing, and the new strain nude rats exist in the obtained F3 generation.
2. The breeding method of nude rats according to claim 1, further comprising the steps of:
(4) After the F3 generation is mature, selecting the male naked rat of the new strain in the F3 generation and the female rat in the F1 generation to carry out cage-closing hybridization until the F4 generation is born;
(5) After the F4 generation is mature, selecting male nude rats in the F3 generation and female rats with the same appearance characteristics as SD rats in the F4 generation for cage crossing, and obtaining the new strain nude rats in the F5 generation.
3. The method for growing nude rats according to claim 1 or 2, wherein the growing is performed in an SPF-class feeding environment at room temperature of 20-26 ℃.
4. A new strain of nude rats obtained by the breeding method according to any one of claims 1 to 3.
5. The new strain of nude rat according to claim 4 for use in medical research.
6. Use of the new strain of nude rat according to claim 4 for the establishment of an immunodeficiency model.
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Citations (5)
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|---|---|---|---|---|
| CN1366813A (en) * | 2002-02-26 | 2002-09-04 | 中国人民解放军第二军医大学 | Method cultivating rat with idiopathic arterial pressure sensitivity reflex function defect |
| US20050022258A1 (en) * | 2003-04-25 | 2005-01-27 | Zheng Cui | Genetically determined mouse model of resistance to transplantable cancers |
| CN107439489A (en) * | 2017-06-15 | 2017-12-08 | 苏州大学附属第二医院 | A kind of method for establishing green fluorescence BALB/c nude mouse models and model application |
| CN108697068A (en) * | 2015-09-17 | 2018-10-23 | 瑞泽恩制药公司 | Selection to the pluripotent cell for generating viable XY female mices |
| CN111418553A (en) * | 2020-04-16 | 2020-07-17 | 段为钢 | Hyperuricemia rat model and construction method thereof |
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Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1366813A (en) * | 2002-02-26 | 2002-09-04 | 中国人民解放军第二军医大学 | Method cultivating rat with idiopathic arterial pressure sensitivity reflex function defect |
| US20050022258A1 (en) * | 2003-04-25 | 2005-01-27 | Zheng Cui | Genetically determined mouse model of resistance to transplantable cancers |
| CN108697068A (en) * | 2015-09-17 | 2018-10-23 | 瑞泽恩制药公司 | Selection to the pluripotent cell for generating viable XY female mices |
| CN107439489A (en) * | 2017-06-15 | 2017-12-08 | 苏州大学附属第二医院 | A kind of method for establishing green fluorescence BALB/c nude mouse models and model application |
| CN111418553A (en) * | 2020-04-16 | 2020-07-17 | 段为钢 | Hyperuricemia rat model and construction method thereof |
Non-Patent Citations (1)
| Title |
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| 赵武述等: "裸鼠与正常鼠杂交子一代的免疫功能研究" * |
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