Kansas State University

  1. K-State home
  2. »1xbet
  3. »1xbet sports betting 7
  4. »ISOLATION OF PLANT GENOMIC 1xbet online sports betting FOR GENOMIC IN SITU HYBRIDIZATION (GISH)

1xbet online games login and ge

Kansas State University
Department of Plant Pathology

4024 Throckmorton
1712 Claflin Road
Manhattan, KS 66506-5502
785-532-6176
785-532-5692 fax
wgrc@k-state.edu

Wheat Genetics Resource Center
Kansas Wheat Innovation Center

1990 Kimball Ave
Manhattan, KS 66502
785-320-4383

ISOLATION OF PLANT GENOMIC 1xbet online sports betting FOR GENOMIC IN SITU HYBRIDIZATION (GISH)

  1. Grind 5 g of plant leaf tissue in liquid nitrogen (do not let the plant material thaw) and pour powder into a 250 1xbet online sports betting flask.
  2. Suspend powdered material in 15 1xbet online sports betting of 2 X CTAB buffer and incubate in a water bath at 65°C for 1 h under slow shaking.
    • 2 X 1xbet online sports betting is 1.4 M NaCl, 100 mM Tris pH 8.0, 2% CTAB (hexadecyltrimethylamonium bromide), 20 mM EDTA, 0.5% Na bisulfide, 1% 2-mercaptoethanol (2-me).
    • When making the CTAB buffer, it is recommended to 1xbet online sports betting the NaCl after the other ingredients except 2-me are in solution. Do not 1xbet online sports betting the 2-me until just before use.
    • The solution without 2-me can be stored at room 1xbet online sports betting .
  3. Let the flasks cool down to room temperature, 1xbet online sports betting 15-mL chloroform/isoamyl alcohol (24:1) and swirl the flask until an emulsion is formed (place the flask on a rotary shaker for 15 min at room temperature).
  4. Put sample into a centrifuge 1xbet online sports betting and spin at 10,000 rpm for 15 min at room temperature.
  5. Transfer the supernatant to a clean tube and add 2/3 volumes of isopropyl alcohol; invert the tubes several times; hook out the precipitated 1xbet online sports betting ; and transfer to a clean 10-mL tube.
  6. Wash the 1xbet online sports betting pellet twice with 70% ethanol. Pour off the ethanol and let the pellet air dry for 1 h.
    add 500 µL of TE and transfer the 1xbet online sports betting into a 1.5-mL tube; add 5 µL RNase A (10 mg/mL) and leave at 4°C overnight or at 37°C for 1 h.
  7. 1xbet online sports betting 500 µL phenol, mix well, and spin for 5 min; transfer the supernatant to another tube.
  8. 1xbet online sports betting 250 µL each of phenol and chloroform, mix well, and spin for 5 min; transfer the supernatant to another tube.
  9. 1xbet online sports betting 500 µL chloroform, mix well, and spin for 5 min; transfer the supernatant to a 10-mL tube.
  10. 1xbet online sports betting TE to a final volume of 3 mL, then 1xbet online sports betting 1/10 volume of 3 M NaAc and 2 volumes of 100% ethanol. Invert the tube several times.
  11. Wash the precipitated 1xbet online sports betting twice with 70% ethanol; transfer the dDNA to a 1.5-mL tube; dry the 1xbet online sports betting pellet; and dissolve into 500 ml TE.
    • As a rule, the cleaner the genomic 1xbet online sports betting , the better the biotin-labeled probe. Therefore, the isolated 1xbet online sports betting may be further purified by using a CsCl-purification procedure. However, the quality of 1xbet online sports betting isolated according to the protocol described above is adequate for GISH in our laboratory.
  12. Determine the 1xbet online sports betting concentration by using spectrophotometer or by electrophoresis on a minigel.