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Nucleic acid probes for identifying haynaldia villosa chromosomes and application thereof

A technology of nucleic acid probes and wheat tufts, applied in the field of chromosome engineering, to achieve the effects of saving time, labor and cost, being fast, accurate and cost-effective, and accelerating the breeding process

Inactive Publication Date: 2017-01-25
SAAS BIOTECH & NUCLEAR TECH RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the oligo probe designed based on a retrotransposon sequence cannot hybridize well in all regions of the chromosome and different chromosomes of the same foreign species like GISH

Method used

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  • Nucleic acid probes for identifying haynaldia villosa chromosomes and application thereof
  • Nucleic acid probes for identifying haynaldia villosa chromosomes and application thereof
  • Nucleic acid probes for identifying haynaldia villosa chromosomes and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Design and preparation of embodiment 1 Oligo sequence probe

[0051] According to the genome sequence of the T. villosa chromosome and the reported repetitive sequences diffusely distributed on the T. villosa chromosome, the 60bp Oligo sequence C1-10-5 and the Oligo sequence C1-10-6 were obtained after screening using BLAST and RepeatMasker software , was synthesized by Invitrogen, and the fluorophore 4-methyl-6-carboxyrhodamine was respectively connected to the 5' ends of the two sequences to obtain Oligo probe C1-10-5 (that is, the present invention Nucleic acid probe 1) and Oligo probe C1-10-6 (ie nucleic acid probe 2 of the present invention). Wherein, the nucleotide sequences of Oligo sequence C1-10-5 and Oligo sequence C1-10-6 from the 5' end to the 3' end are shown in SEQ ID NO.1 and SEQ ID NO.2 in the sequence table.

Embodiment 2

[0052] Fluorescence in situ hybridization verification experiment of embodiment 2 tuft wheat chromosome

[0053] (1), get the root of the durum wheat-Tutritus villosa amphidiploid TH1W (2n=42, AABBVV):

[0054] 1) Put the seeds on wet filter paper and incubate at 23°C for 24h.

[0055] 2) Transfer the seeds to culture at 4°C for 24 hours.

[0056] 3) Transfer the seeds to culture at 23°C until they root.

[0057] 4) When the root length reaches 1-2 cm, take the root and place it in a moistened 0.5ml EP tube (the EP tube is placed on ice). The 0.5ml EP tube should be poked a small hole with a diameter of about 3mm in advance to facilitate the treatment of the root tip with nitrous oxide.

[0058] 5) Put the EP tube with the root into the nitrous oxide tank, and pass the nitrous oxide (0.9Mpa) through the nitrous oxide tank for 2 hours.

[0059] 6) Take out the EP tube and place it on ice, add 90% acetic acid into it, and fix the root tip for 8-10 min. with ddH 2 O wash th...

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Abstract

The invention provides a pair of oligonucleotides and nucleic acid probes prepared from the same. The two nucleic acid probes can form better dispersive distribution on most of haynaldia villosa chromosomes, and by combining the nucleic acid probes with known oligomeric nucleic acid probes Oligo-pTa535, the haynaldia villosa chromosomes and / or chromosome segments in the genetic background of wheat can be identified quickly and accurately. The technology has the advantages of time saving, labor saving, low cost and capability of reaching a similar GISH (genomic in situ hybridization) effect.

Description

technical field [0001] The invention belongs to the field of chromosome engineering, and in particular relates to a nucleic acid probe for identifying the chromosome of T. villosa and its application. Background technique [0002] The annual diploid Triticum tufts (2n=14, VV) is an important relative of wheat, carrying a variety of excellent genes for insect resistance, disease resistance and abiotic stress resistance (De Pace C 2011). As early as the 1950s, researchers began to conduct studies on the transfer of the genetic material of the annual diploid T. villosa to the genetic background of wheat. Sears (1953) first used emmer wheat to cross with Triticum triticum to obtain durum wheat-Custicum triticum amphidiploid plants. Blanco et al. (1988) obtained a double-end addition line of 6VS in durum wheat, which was genetically stable and resistant to powdery mildew. In 1991, Blanco et al. identified the complete monomer addition lines of durum wheat "Creso" and wheat tuft...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6841C12Q1/6895C12Q2563/107
Inventor 张洁蒋云宣朴郭元林
Owner SAAS BIOTECH & NUCLEAR TECH RES INST
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