Preparation method and detection of hybridization probe and gene chip for main strains of plum pox virus
A technology of gene chip and plum pox virus, which is applied in the field of preparation and detection of hybridization probes and gene chips of main strains of plum pox virus, can solve the problem that there are no reports on the detection and identification of different strains of plum pox virus, etc.
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Embodiment 1
[0078] A hybridization probe for main strains of plum pox virus, comprising:
[0079] PPV-D: 5'-CAACAAAACCAGTTTCACAGGTGCCAGGACCTCAACTGCAA-3'
[0080] PPV-M: 5'-CGGTAAGACCAGTTCCTCCAATTTCAGGGACCAAACCGCGG-3'
[0081] PPV-EA: 5'-CCACTAGGACAGTGCCTCACACAACAACTACTACACCTCCT-3'
[0082] PPV-Rec: 5'-CGATAAGACCAGTTTCTCCAATTTCAGGGGCCACACCGCAG-3'
[0083] PPV-W: 5'-GCGTGAAGCCAACTACATCAGCAACAATTAACCCAACGTCT-3'
[0084] PPV-C: 5'-ATGTGAGACCGATTGCACCAGTAGTGACAAGTCCATTCTCG-3'.
Embodiment 2
[0086] A method for preparing a gene chip, comprising the following steps:
[0087] A. Preparation of gene chip spotting:
[0088] Take 5ul of purified probe, add 5ul of spotting solution and mix well, the probe concentration is 10μm; add it to the sample spotter, spot the oligonucleotide probe on the aldehyde-based glass slide, and pass through the amino group at the 5' end of the probe carry out covalent immobilization;
[0089] Wherein said probes include:
[0090] PPV-D: 5'-CAACAAAACCAGTTTCACAGGTGCCAGGACCTCAACTGCAA-3'
[0091] PPV-M: 5'-CGGTAAGACCAGTTCCTCCAATTTCAGGGACCAAACCGCGG-3'
[0092]PPV-EA: 5'-CCACTAGGACAGTGCCTCACACAACAACTACTACACCTCCT-3'
[0093] PPV-Rec: 5'-CGATAAGACCAGTTTCTCCAATTTCAGGGGCCACACCGCAG-3'
[0094] PPV-W: 5'-GCGTGAAGCCAACTACATCAGCAACAATTAACCCAACGTCT-3'
[0095] PPV-C: 5'-ATGTGAGACCGATTGCACCAGTAGTGACAAGTCCATTCTCG-3';
[0096] B. PCR amplification and sample labeling:
[0097] B1. Extraction of total RNA from susceptible plants;
[0098] Weigh 0.1...
Embodiment 3
[0117] A method for preparing a gene chip, comprising the following steps:
[0118] A. Preparation of gene chip spotting:
[0119] Take 5ul of purified probe, add 5ul of spotting solution and mix well, the probe concentration is 10μm; add it to the sample spotter, spot the oligonucleotide probe on the aldehyde-based glass slide, and pass through the amino group at the 5' end of the probe carry out covalent immobilization;
[0120] Wherein said probes include:
[0121] PPV-D: 5'-CAACAAAACCAGTTTCACAGGTGCCAGGACCTCAACTGCAA-3'
[0122] PPV-M: 5'-CGGTAAGACCAGTTCCTCCAATTTCAGGGACCAAACCGCGG-3'
[0123] PPV-EA: 5'-CCACTAGGACAGTGCCTCACACAACAACTACTACACCTCCT-3'
[0124] PPV-Rec: 5'-CGATAAGACCAGTTTCTCCAATTTCAGGGGCCACACCGCAG-3'
[0125] PPV-W: 5'-GCGTGAAGCCAACTACATCAGCAACAATTAACCCAACGTCT-3'
[0126] PPV-C: 5'-ATGTGAGACCGATTGCACCAGTAGTGACAAGTCCATTCTCG-3';
[0127] The specific method of spotting: use a spotting machine to spot samples in an array arrangement, the center distance betwee...
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