CN110656127A - Method for efficiently obtaining transgenic radish - Google Patents
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Abstract
The invention discloses a method for efficiently obtaining transgenic radish. The method comprises the following steps: (1) preparing a flower soaking material; (2) constructing and purifying pCAMBIA3301-eGFP carrier plasmid; (3) preparing agrobacterium; (4) dip dyeing; (5) and (5) harvesting the seeds. The invention ensures that the operation of obtaining the transgenic plant of the radish is simpler and more convenient, a complex and unstable radish regeneration system is not needed, the time and money spent on tissue culture of the radish are reduced, and the experiment cost is reduced by more than 30%; meanwhile, the genetic transformation efficiency of the radish is increased to about 1% from less than 0.2% of that of the traditional method, and is improved by about 5 times.
Description
Technical Field
The invention belongs to the technical field of genetic engineering, and particularly relates to a method for efficiently obtaining transgenic radish.
Background
Radish (Raphanus sativus L.) is an important cruciferous vegetable crop in China, the annual planting area is about 120 million hectares, the radish is located in the third place of the country, and the radish plays an important role in vegetable production and supply. In the aspect of basic research, the development of radish research and development is extremely inconsistent with the status and importance of radish cultivation and production. Particularly, in the aspect of molecular biology, although the whole genome sequencing work of radish has been completed by japanese scientists in 2014 and two more complete whole genome databases have been published successively in 2015 (japan) and 2016 (korea), many experiments such as gene function verification and the like can only be performed on arabidopsis thaliana belonging to the same family of brassicaceae due to low genetic transformation efficiency and complicated and unstable operation of a regeneration system, and the verification result cannot truly reflect the situation of radish, and leading edge science technologies such as CRISPR gene editing and the like cannot be performed on radish successfully so far. For the reasons, the overall level of the basic research of radishes is always in the middle-lower level, and a periodical with higher influence factor (IF >5.0) is published for a few times. The radish transformation rate is low, and the transgenic plant is difficult to obtain, so that the radish transformation rate is one of the main reasons for seriously delaying the research progress of radish in the aspect of molecular biology.
The agrobacterium-mediated floral dip genetic transformation method is simpler and more convenient in the existing plant genetic transformation method independent of tissue culture. The method of soaking flower is used for transgenosis, and has the advantages of avoiding tissue culture and regeneration process and directly obtaining transgenic T0The seeds are used for generation, and compared with other methods without tissue culture, the method has lower requirements on instruments, equipment and technical operation. Therefore, the floral dip method is suitable for tissue culture and in vitro regeneration of plants which are difficult. The research history of the flower soaking method is long, experiments in arabidopsis thaliana of cruciferae are successful and reported in 1987, and the genetic transformation of the flower soaking method can be carried out in brassica napus of the same genus brassica. However, since radish is a typical cross-pollinated plant, most materials have strong self-incompatibility, and can not be self-fertilized or have little fruit when being directly pollinated, the agrobacterium-mediated flower soaking genetic transformation method is not reported in radish genetic transformation research.
Disclosure of Invention
Aiming at the defects in the prior art, the invention provides a method for efficiently obtaining transgenic radish, which can obtain a stable radish regeneration system.
In order to achieve the purpose, the technical scheme adopted by the invention for solving the technical problems is as follows:
a method for efficiently obtaining transgenic radish comprises the following steps:
(1) preparation of flower-soaking Material
Culturing the radish after budding for 5-6 days for 40-50 days, pulling out the radish, removing leaves above stem tip growing points, storing at 4-8 ℃ for 20-30 days, screening out the radish without diseases, planting under 3000-5000 lx illumination, culturing until bolting and blooming, and removing bloomed flowers for later use;
(2) constructing and purifying pCAMBIA3301-eGFP carrier plasmid; wherein the nucleotide sequence of the eGFP gene is shown as SEQ ID NO. 1;
(3) preparation of Agrobacterium
Transforming the purified pCAMBIA3301-eGFP vector plasmid into competent cells, carrying out colony identification, and storing at-80 to-60 ℃;
(4) dip dyeing
Taking out the radish plants preserved in the step (1), putting the radish plants into a dip dyeing solution for dip dyeing for 1-5 min after flowers on the radish plants bloom, bagging the flowers, carrying out shading cultivation for 5-7 days, then dip dyeing the flowers for 1-5 min again, and repeating for 3-5 times;
(5) harvesting seeds
And (5) after the dip dyeing in the step (4) is finished for 30-40 days, harvesting after the seeds are mature.
Further, the radish in the step (1) is a radish with self-mating fruiting capacity.
Furthermore, the radish is purple-flower radish.
Further, the construction process of the vector plasmid in the step (2) is as follows:
(1) respectively extracting and purifying plasmids pCAMBIA3301 and pUC57-eGFP, and then respectively carrying out double enzyme digestion reaction on the plasmids pCAMBIA3301 and pUC57-eGFP by adopting HindIII and Sac I;
(2) after being warmed at 37 ℃ for 12h, the large fragment of the plasmid pCAMBIA3301 and the small fragment of the plasmid pUC57-eGFP are recovered by electrophoresis respectively;
(3) the fragments recovered in step (2) were ligated with T4 ligase and reacted at 16 ℃ for 12 hours to obtain pCAMBIA3301-eGFP vector plasmid.
Further, the competent cell in step (3) is Agrobacterium GV 3101.
Further, the preparation method of the dip dyeing solution in the step (4) comprises the following steps:
(1) taking out the agrobacterium liquid frozen in the step (3), streaking on a resistant LB solid culture medium plate, inverting the plate in a 28 ℃ incubator for 48 hours, picking out a single colony, adding the single colony into a resistant LB liquid culture medium, and performing shake culture at 28 ℃ and 200r/min for 12 hours;
(2) after the culture is finished, transferring the strain into resistant LB liquid culture, and continuing shaking culture at 28 ℃ and 200r/min for 12-16 h until the OD value of the strain liquid is 0.6-0.8, and taking out the strain liquid;
(3) and then centrifuging at 5000rpm for 10-15 min, collecting the thalli, and then resuspending the thalli by using an MS liquid culture medium containing 5% of sucrose and 0.1% of silwet.
Further, the resistant LB solid medium included 50. mu.g/mL kanamycin, 50. mu.g/mL rifampicin, and 50. mu.g/mL gentamicin.
Further, the resistant LB liquid medium in step (1) includes 50. mu.g/mL kanamycin, 50. mu.g/mL rifampicin, and 50. mu.g/mL gentamicin.
Further, the resistant LB liquid medium in step (2) includes 50. mu.g/mL kanamycin, 25. mu.g/mL rifampicin, and 50. mu.g/mL gentamicin.
The invention has the beneficial effects that:
1. the invention ensures that the operation of obtaining the transgenic plant of the radish is simpler and more convenient, a complex and unstable radish regeneration system is not needed, the time and money spent on tissue culture of the radish are reduced, and the experiment cost is reduced by more than 30%; meanwhile, the genetic transformation efficiency of the radish is increased to about 1% from less than 0.2% of that of the traditional method, and is improved by about 5 times.
2. The invention develops scientific research experiments in the aspect of radish gene function verification, can effectively break the research barrier of CRISPR (gene editing) technology, and promotes the research process of radish molecular biology and molecular breeding.
Drawings
FIG. 1 shows the construction process of pCAMBIA3301-eGFP vector plasmid;
FIG. 2 shows the results of fluorescent detection of eGFP gene in radish seedlings;
FIG. 3 shows the result of electrophoresis detection of the expression of eGFP gene by RT-PCR.
Detailed Description
The following description of the embodiments of the present invention is provided to facilitate the understanding of the present invention by those skilled in the art, but it should be understood that the present invention is not limited to the scope of the embodiments, and it will be apparent to those skilled in the art that various changes may be made without departing from the spirit and scope of the invention as defined and defined in the appended claims, and all matters produced by the invention using the inventive concept are protected.
Example 1
A method for efficiently obtaining transgenic radish comprises the following steps:
carrying out artificial germination on purple-flower radish seeds or other radish seeds with good selfing and fructification capacity, and sowing the seeds in the field after 5 days of germination. After about 40 days of vegetative growth in a natural environment, the radish is pulled out, most of leaves above the growing point of the stem tip are removed, and the radish is moved into a refrigeration house at the temperature of 4-8 ℃ and placed for 20 days. Then transplanting the purple-flower radishes which are not damaged into flower pots with the diameter of about 25cm, and transplanting one radish in each pot. And then moving the flowerpot to a field natural light or indoor light supplement environment (the illumination intensity is more than 3000lx), carrying out reproductive growth for about 30 days, bolting and flowering, removing all bloomed flowers from the period from the initial flowering to the full-bloom period, and placing the bloomed flowers indoors for later use.
(2) Construction of pCAMBIA3301-eGFP vector plasmid (shown in FIG. 1)
a. Extracting and purifying plasmids pCAMBIA3301 and pUC57-eGFP (connected with 35S promoter and Nos terminator) by using a Tiangen (TIANGEN) plasmid extraction kit; the sequence of the eGFP gene (GenBank: NC-025025.1, 720bp) is shown in SEQ ID NO. 1;
b. HindIII and Sac I are adopted to respectively carry out double enzyme digestion reaction on plasmid pCAMBIA3301 and pUC 57-eGFP;
c. after 12h of warm bath at 37 ℃, the large fragment of the plasmid pCAMBIA3301 and the small fragment of the plasmid pUC57-eGFP are respectively recovered by electrophoresis by using a common agarose gel DNA recovery kit (centrifugal column type) of Tiangen (TIANGEN);
d. the large and small fragments were recovered by ligation with T4 ligase and reacted at 16 ℃ for 12 hours to give pCAMBIA3301-eGFP vector plasmid, which was purified.
(3) Preparation of Agrobacterium
The purified pCAMBIA3301-eGFP vector plasmid is transformed into agrobacterium CV3101 competent cells by an electric shock transformation method, plate coating identification is carried out, and then the bacterial liquid is stored in a refrigerator at the temperature of 80 ℃ below zero for later use.
(4) Preparation of the Dip dyeing liquor
a. Taking out the frozen agrobacterium liquid obtained in the step (3), streaking on a resistant LB solid culture medium plate, inverting the plate in an incubator at 28 ℃ for culturing for 48 hours, picking out a single colony, adding the single colony into a 50mL standard centrifuge tube filled with 20mL of resistant LB liquid culture medium, and culturing for 12 hours at 28 ℃ and 200r/min with small shaking; the resistant LB solid culture medium comprises 50 mug/mL kanamycin, 50 mug/mL rifampicin and 50 mug/mL gentamicin; the resistant LB liquid culture medium comprises 50 mug/mL kanamycin, 50 mug/mL rifampicin and 50 mug/mL gentamicin;
b. after the culture is finished, transferring the strain into a 500mL triangular flask filled with 200mL of resistant LB liquid culture medium, and culturing the strain for 12-16 h at 28 ℃ under a large shaking at 200r/min until the OD value of the strain liquid is 0.6-0.8, and taking out the strain liquid; the resistant LB liquid culture medium comprises 50 mug/mL kanamycin, 25 mug/mL rifampicin and 50 mug/mL gentamicin;
c. then centrifuging at 5000rpm for 10min, collecting thallus, and then resuspending the thallus with MS liquid culture medium containing 5% sucrose and 0.1% silwet.
(5) Dip dyeing
Adding 100mL of dip dyeing solution into a beaker with the specification of 250mL, taking out the radish plant stored in the step (1), placing the radish plant into the dip dyeing solution for dip dyeing for 1min after flowers on the radish plant bloom, leaving the uninvaded flowering branch of the same plant as a control, and bagging and marking adjacent flowering branches. After shading treatment for 24 hours, the cap bag was removed. After 7 days, the newly bloomed flowers were dip-dyed again and repeated 3 times.
(6) Harvesting seeds
And after dip dyeing for 30-40 days, respectively collecting seeds according to marks after the seed pods turn yellow and the seeds are mature, naturally drying, threshing and storing.
Example 2 validation of transformation efficiency
Will obtain T0And (5) seed generation, sowing the seeds in a 128-hole seedling tray, spraying a Basta solution with the content of 0.225g/Lppt, and continuously managing the obtained resistant seedlings. Meanwhile, the control seeds of the same strain which are not subjected to flower soaking treatment are sown for subsequent identification.
Taking out the resistant seedlings, washing and placing under a fluorescence confocal microscope. Under the condition of fluorescence excitation, the condition of the root tip and root hair part cells is observed, the green fluorescence of the eGFP gene can be observed in the positive seedlings (figure 2), and the result is recorded. Statistical analysis gave a conversion efficiency of 0.96%.
Meanwhile, the RNA of the resistant seedling leaf is extracted, RT-PCR (reverse transcription-polymerase chain reaction) is carried out, the non-impregnated plant and the plant reference Actin gene (GenBank: KC751568.1, 296bp, the sequence is shown in SEQ ID NO.2) are set as the reference, the Actin1, the Actin2, the eGFP1 and the eGFP2 are respectively used as the primers (the sequence is shown in the table 1 in detail), the PCR amplification is carried out, the expression condition of the target gene is verified (shown in the table 3), the amplification reaction system is shown in the table 2, and the amplification program is shown in the table 3.
TABLE 1 RT-PCR primers
TABLE 2 PCR reaction System
TABLE 3 PCR amplification procedure
As shown in FIG. 3, a in FIG. 3 is the expression of eGFP gene; b is the expression condition of an Actin gene; the strip MII is a Tiangen (TIANGEN) MarkerII product, the strip ck-is a control without flower soaking treatment, the strip ck + is a plasmid containing a corresponding gene, and the strips 1-6 are partial resistant seedlings obtained after flower soaking treatment; meanwhile, the calculation result shows that the transformation efficiency is 1.15%, which is basically consistent with the result verified by a fluorescence confocal microscope, and shows that the pCAMBIA3301-eGFP vector plasmid is successfully transformed into radish seeds to obtain transgenic plants.
Sequence listing
<110> research institute for sorghum oryzae, academy of agricultural sciences, Sichuan province
<120> a method for efficiently obtaining transgenic radish
<141> 2019-11-06
<160> 6
<170> SIPOSequenceListing 1.0
<210> 1
<211> 720
<212> DNA
<213> radish (Raphanus sativus)
<400> 1
atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac 60
ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac 120
ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc 180
ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag 240
cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc 300
ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg 360
gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac 420
aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac 480
ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc 540
gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac 600
tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc 660
ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa 720
<210> 2
<211> 296
<212> DNA
<213> radish (Raphanus sativus)
<400> 2
agctgcaggg atccacgaga cgacctacaa ctcgatcatg aagtgtgacg tggatatcag 60
gaaggacttg tacggtaaca ttgtgctcag tggtggaacc actatgttct cgggtattgc 120
agaccgtatg agcaaagaga tcacagcact tgcaccaagc agcatgaaga ttaaggtcgt 180
tgcacctccg gagaggaagt acagtgtctg gatcggtggt tccattcttg cttccctcag 240
cacattccag cagatgtgga tctccaaggc tgagtatgat gaagcaggtc caggca 296
<210> 3
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<213> Artificial Sequence (Artificial Sequence)
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agctgcaggg atccacgaga cgac 24
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<213> Artificial Sequence (Artificial Sequence)
<400> 4
tgcctggacc tgcttcatca tactc 25
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<213> Artificial Sequence (Artificial Sequence)
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atggtgagca agggcgagga gc 22
<210> 6
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<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 6
ttacttgtac agctcgtcca tgcc 24
Claims (8)
1. A method for efficiently obtaining transgenic radish is characterized by comprising the following steps:
(1) preparation of flower-soaking Material
Culturing the radish after budding for 5-6 days for 40-50 days, pulling out the radish, removing leaves above stem tip growing points, storing at 4-8 ℃ for 20-30 days, screening out the radish without diseases, planting under 3000-5000 lx illumination, culturing until bolting and blooming, and removing bloomed flowers for later use;
(2) constructing and purifying pCAMBIA3301-eGFP carrier plasmid;
(3) preparation of Agrobacterium
Transforming the purified pCAMBIA3301-eGFP vector plasmid into competent cells, carrying out colony identification, and storing at-80 to-60 ℃;
(4) dip dyeing
Taking out the radish plants preserved in the step (1), putting the radish plants into a dip dyeing solution for dip dyeing for 1-5 min after flowers on the radish plants bloom, bagging the flowers, carrying out shading cultivation for 5-7 days, then dip dyeing the flowers for 1-5 min again, and repeating for 3-5 times;
(5) harvesting seeds
And (5) after the dip dyeing in the step (4) is finished for 30-40 days, harvesting after the seeds are mature.
2. The method for efficiently obtaining transgenic radish according to claim 1, wherein the radish in step (1) is self-fruiting radish.
3. The method for efficiently obtaining transgenic radish according to claim 1, wherein the vector plasmid in step (2) is constructed by the following steps:
(1) respectively extracting and purifying plasmids pCAMBIA3301 and pUC57-eGFP, and then respectively carrying out double enzyme digestion reaction on the plasmids pCAMBIA3301 and pUC57-eGFP by adopting HindIII and Sac I;
(2) after being warmed at 37 ℃ for 12h, the large fragment of the plasmid pCAMBIA3301 and the small fragment of the plasmid pUC57-eGFP are recovered by electrophoresis respectively;
(3) the fragments recovered in step (2) were ligated with T4 ligase and reacted at 16 ℃ for 12 hours to obtain pCAMBIA3301-eGFP vector plasmid.
4. The method for efficiently obtaining transgenic radish according to claim 1, wherein the competent cell in step (3) is Agrobacterium GV 3101.
5. The method for efficiently obtaining transgenic radish according to claim 1, wherein the preparation method of the dip dyeing solution in the step (4) is as follows:
(1) taking out the agrobacterium liquid frozen in the step (3), streaking on a resistant LB solid culture medium plate, inverting the plate in a 28 ℃ incubator for 48 hours, picking out a single colony, adding the single colony into a resistant LB liquid culture medium, and performing shake culture at 28 ℃ and 200r/min for 12 hours;
(2) after the culture is finished, transferring the strain into resistant LB liquid culture, and continuing shaking culture at 28 ℃ and 200r/min for 12-16 h until the OD value of the strain liquid is 0.6-0.8, and taking out the strain liquid;
(3) and then centrifuging at 5000rpm for 10-15 min, collecting the thalli, and then resuspending the thalli by using an MS liquid culture medium containing 5% of sucrose and 0.1% of silwet.
6. The method for efficiently obtaining transgenic radish according to claim 5, wherein the resistant LB solid medium comprises 50 μ g/mL kanamycin, 50 μ g/mL rifampicin and 50 μ g/mL gentamicin.
7. The method for efficiently obtaining transgenic radish according to claim 5, wherein the resistant LB liquid medium in step (1) comprises 50 μ g/mL kanamycin, 50 μ g/mL rifampicin and 50 μ g/mL gentamicin.
8. The method for efficiently obtaining transgenic radish according to claim 5, wherein the resistant LB liquid medium in step (2) comprises 50 μ g/mL kanamycin, 25 μ g/mL rifampicin and 50 μ g/mL gentamicin.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110295174A (en) * | 2019-03-13 | 2019-10-01 | 济宁学院 | The transgenic arabidopsis strain and its construction method of FIPV gene overexpression |
| CN110295179A (en) * | 2019-08-05 | 2019-10-01 | 浙江大学 | Turnip disease-resistant related gene BrPGIP8 and its application |
-
2019
- 2019-11-06 CN CN201911077540.9A patent/CN110656127A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110295174A (en) * | 2019-03-13 | 2019-10-01 | 济宁学院 | The transgenic arabidopsis strain and its construction method of FIPV gene overexpression |
| CN110295179A (en) * | 2019-08-05 | 2019-10-01 | 浙江大学 | Turnip disease-resistant related gene BrPGIP8 and its application |
Non-Patent Citations (3)
| Title |
|---|
| IAN S CURTIS等: "Expression of an antisense GIGANTEA (GI) gene fragment in transgenic radish causes delayed bolting and flowering", 《TRANSGENIC RES》 * |
| 周俊国等: "《园艺植物育种技术》", 31 August 2006, 中国农业出版社 * |
| 喻晓敏等: "根癌农杆菌介导的萝卜遗传转化方法研究", 《北方园艺》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114958904A (en) * | 2022-05-20 | 2022-08-30 | 南京农业大学 | Method for rapidly obtaining radish transgenic material |
| CN114958904B (en) * | 2022-05-20 | 2023-10-24 | 南京农业大学 | Method for rapidly obtaining radish transgenic material |
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