A method for developing ramie genome ssr markers in large quantities and the primers developed therefor
A genome, large-scale technology, applied in the field of genetic engineering, to achieve the effect of high cost, high efficiency and low cost
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Embodiment 1
[0022] (1) Determine the enzyme digestion plan, gel cutting range, sequencing volume, etc. to ensure the density and uniformity of molecular marker development to ensure that the expected experimental purpose is achieved.
[0023] The DNA of Ramie No.1 and Hejiang Qingma was extracted.
[0024] Enzyme digestion scheme selection
[0025] According to the selection principle of closely related species, the cannabis genome was finally selected as the reference genome for enzyme digestion prediction.
[0026] Ramie species information and related species information are as follows:
[0027] Ramie information: Genome size is 716Mb, GC content is 49%;
[0028] Related species cannabis information: Genome size is 757Mb, GC content is 24%, download link:
[0029] ftp: / / ftp.ncbi.nlm.nih.gov / genbank / genomes / Eukaryotes / plants / Cannabis_sativa / canSat3 / .
[0030] Candidate Digestion Protocol Information
[0031] The cannabis genome was systematically analyzed using enzyme digesti...
Embodiment 2
[0058] Primer polymorphism detection
[0059] 1 Materials and methods
[0060] 1.1 Materials
[0061] 24 selfed offspring of Zhongzhu 1;
[0062] There are 24 ramie varieties, and the variety names are shown in Table 4.
[0063]
[0064] 1.2 Method
[0065] 1.2.1 Genomic DNA extraction
[0066] DNA was extracted from 24 selfed offspring of Zhongzhu No. 1 and new shoots from 24 individual plants of ramie varieties using the Tiangen kit. After the concentration of the extracted DNA is detected by electrophoresis, the concentration of the sample DNA is calculated and diluted to the required concentration.
[0067] 1.2.2 PCR using the designed SSR primers
[0068] PCR reaction system: 20μL reaction system composition is shown in Table 5
[0069] Table 5 PCR reaction system composition
[0070] System composition
Final concentration
Mg 2+
2.0mmol / L
Taq Buffer
1×
dNTP Mix
200μmol / L each
Taq Enzymes
1U
Primers...
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