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Method for identifying indica rice and japonica rice by inserting or deleting molecular marker in rice DNA

A technology of molecular markers and DNA sequences, applied in the field of identification of indica and japonica rice, can solve the problems that affect the efficiency of indica and japonica rice identification, time-consuming, inaccurate detection results, etc.

Inactive Publication Date: 2008-12-17
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The biggest disadvantage of this method is: first, because the morphological traits are greatly affected by the environment, it is easy to cause inaccurate detection results; second, the morphological measurement must be planted with the tested rice sample, and it usually takes 3-5 months from sowing to maturity. It takes a long time; third, morphological measurement requires a certain sample population, so more rice samples are required; this greatly affects the efficiency of identification of indica and japonica rice

Method used

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  • Method for identifying indica rice and japonica rice by inserting or deleting molecular marker in rice DNA
  • Method for identifying indica rice and japonica rice by inserting or deleting molecular marker in rice DNA
  • Method for identifying indica rice and japonica rice by inserting or deleting molecular marker in rice DNA

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] InDel molecular markers were used to identify 10 typical indica and 10 typical japonica rice varieties determined by "Cheng's index method".

[0044] Table 1. Embodiment 1 identification experimental material:

[0045]

[0046]

[0047] The reaction system used is as follows:

[0048] The PCR reaction conditions and procedures are as follows: (20 μl system):

[0049] 1) 10×PCR buffer (containing 25mM MgCl 2 ) 2μl

[0050] 2) 2.5mM dNTP Mixture 1.6μl

[0051] 3) 1μl of 10μM PCR primers

[0052] 4) 1U Taq DNA Polymerase 0.2μl

[0053] 5) DNA template 20ng

[0054] 6)ddH 2 O (double distilled water) to make up to 20μl

[0055] PCR reaction program:

[0056] 1) Pre-denaturation at 94°C for 4 minutes

[0057] 2) Denaturation at 94°C for 40 seconds

[0058] 3) Annealing temperature: 52-58°C, annealing time: 30-45 seconds

[0059] 4) Extend at 72°C for 40 seconds

[0060] 5) The cycle steps are 2) 3) 4), 36 cycles

[0061] 6) Extend at 72°C for 10 minutes

...

Embodiment 2

[0068] Identification of rice varieties with unknown indica and japonica characteristics using InDel molecular markers. Twenty samples were randomly selected from China, Bangladesh, India, Japan, Laos, Thailand and Malaysia for identification of indica or japonica rice.

[0069] Table 2. Embodiment 2 identification experimental materials:

[0070]

[0071]

[0072] The reaction system used is as follows:

[0073] The PCR reaction conditions and procedures are as follows: (20 μl system):

[0074] 1) 10×PCR buffer (containing 25mM MgCl 2 ) 2μl

[0075] 2) 2.5mM dNTP Mixture 1.6μl

[0076] 3) 10μM PCR primer 1μl

[0077] 4) 1U Taq DNA Polymerase 0.2μl

[0078] 5) DNA template 20ng

[0079] 6)ddH 2 O (double distilled water) to make up to 20μl

[0080] PCR reaction program:

[0081] 1) Pre-denaturation at 94°C for 4 minutes

[0082] 2) Denaturation at 94°C for 40 seconds

[0083] 3) Annealing temperature: 52-58°C, annealing time: 30-45 seconds

[0084] 4) Extend...

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Abstract

The invention pertains to the technical field of biological discrimination, in particular to a method for discriminating indica rice from japonica rice by utilizing molecular markers of rice DNA insertion or deletion. The method obtains 34 pairs of differential DNA primers designed by InDel differential fragment by making a comparison of whole genome sequence between indica rice cultivar 93-11 and japonica rice cultivar (Nipponbare). The DNA extraction, the amplification and ionophortic separation of DNA fragments and the analysis and calculation of an electrophoretogram of a target rice cultivar are carried out to discriminate indica rice from japonica rice. Precisely speaking, the 34 pairs of InDel primer groups are utilized and analysis and calculation are carried out according to a finger-print obtained on the basis of a polymerase chain reaction and vertical slab gel electrophoresis; the characteristics of indica rice or japonica rice of rice samples detected are worked out according to the average frequency (Fi or Fj) showing on indica rice alleles and japonica rice alleles on 34 InDel locuses. The invention has the advantages of simple, fast and convenient method, few samples for detection and correct discrimination results, thus having good prospect of promotion and application.

Description

technical field [0001] The invention belongs to the technical field of biological identification, and in particular relates to a method for identifying indica rice and japonica rice. technical background [0002] Indica and japonica are the two most important genetically differentiated groups of Asian cultivated rice (Oryza sativa L.). There has been obvious reproductive isolation between typical indica rice and typical japonica rice, so indica rice and japonica rice are usually treated as two subspecies in classification, namely: indica rice subspecies (O. sativa subsp. indica) and japonica rice subspecies (O. sativa subsp. iaponica). Indica rice and japonica rice have produced large differences in morphology, physiological and biochemical traits and quality, and also have differences in adaptability to different ecological environments. Indica rice is suitable for low-latitude and low-altitude tropical and subtropical environments, while japonica rice It is more suitable...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 卢宝荣蔡星星
Owner FUDAN UNIV
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