Molecule labeling method for female gene of jie gourd

Molecule labeling method for female gene of jie gourd

  • CN 1,452,857 B
  • Filed: 05/16/2003
  • Issued: 04/14/2010
  • Est. Priority Date: 05/16/2003
  • Status: Active Grant
First Claim
Patent Images

1. a molecule marking method that saves the melon female gene is characterized in that it comprises the steps:

  • The acquisition and the maintenance of the full female plant of the first step system;

    through selfing for many years, orthoselection selects the full female plant of joint melon from the strong female plant of joint melon, and full female plant sprays AgNO at 3~

    5 leaf periods 3, induce the generation male flower, carry out selfing then, be seed thereby obtain the full female plant of joint melon, and then make the full female plant of joint melon system be maintained and continue;

    Second goes on foot acquisition and the maintenance that full staminiferous plant is;

    through selfing for many years, orthoselection selects the full staminiferous plant of joint melon from the strong staminiferous plant of joint melon, and the male flower on the full staminiferous plant and its are hybridized with the female flower on the strong staminiferous plant of relationship, thereby acquisition saves the melon cenospecies, from F 1In separablely after the selfing go out full staminiferous plant, and then make the full staminiferous plant of joint melon system be maintained and continue;

    Full female plant of the 3rd step, the acquisition of staminiferous plant entirely;

    utilizing the full female plant of the joint melon that isozygotys system is to hybridize with full staminiferous plant, obtains F 1Seed, plantation F 1Seed can obtain the F of segregating generation 2Plant therefrom can filter out full female plant, full staminiferous plant;

    The acquisition of full female plant DNA of the 4th step, full staminiferous plant DNA;

    get the full female plant of 7~

    10 strains, full staminiferous plant respectively, composite samples extracts full female plant, full staminiferous plant DNA by the CTAB method;

    The 5th step molecule marker reaction system;

    RAPD reacts cumulative volume 25 μ

    l, contains the MgCl of 25mmol/L 2, the dNTP0.25 μ

    l of 10mmol/L, Taq archaeal dna polymerase 0.5U, 10 base random primers, 1 μ

    l, template DNA 20~

    30ng, the thermal cycle conditions of amplified reaction;

    94 ℃

    , 1min;

    36 ℃

    , 2min;

    72 ℃

    , 1min;

    Change 94 ℃

    into after 5 circulations, 0.5min;

    36 ℃

    , 1min;

    72 ℃

    , 1.5min;

    Extend 10min in 72 ℃

    after 35 circulations;

    Get 8~

    10 μ

    l reaction product, with 2 μ

    l tetrabromophenol sulfonphthalein mixings, click and enter in 1.0% sepharose that contains 0.5mg/L EB, electrophoresis is 2~

    3 hours under 5V/cm strength of electric field, takes out gel, adopt ultraviolet gel analysis system to analyze, filter out 1 primer;

    OPQ-45 from 198 random primers, its sequence is TGAGCGGACA, can amplify a special band in complete female gene pool, the size of this amplified band is 1150bp, called after OPQ-45 1150

View all claims
    ×
    ×

    Thank you for your feedback

    ×
    ×