Oligonucleotide prehybridization solution

0.05 M sodium phosphate (pH 6.8)

5x Denhardt's solution

100 pg/ml denatured, fragmented salmon sperm DNA Phosphate Buffers

Gomori buffers, the most commonly used phosphate buffers, consist of a mixture of monobasic dihydrogen phosphate and dibasic monohydrogen phosphate. By varying the amount of each salt, a range of buffers can be prepared that buffer well between pH 5.8 and pH 8.0 (please see the tables below). Phosphates have a very high buffering capacity and are highly soluble in water. However, they have a number of potential disadvantages:

Phosphates inhibit many enzymatic reactions and procedures that are

• the foundation of molecular cloning, including cleavage of DNA by many restriction enzymes, ligation of DNA, and bacterial transformation.

Because phosphates precipitate in ethanol, it is not possible to

• precipitate DNA and RNA from buffers that contain significant quantities of phosphate ions.

• Phosphates sequester divalent cations such as Ca2+ and Mg2+.

Preparation of 0.1 M Potassium Phosphate Buffer at 25 DC

pH

Volume of 1 m K,HP04 (ml)

Volume of 1 m KH.PO, (ml)

sa

8.5

91.5

S.0

13.2

86.8

6.2

19.2

80.8

G.4

27.8

72.2

6.6

38.1

Gl,9

S.8

49.7

50.3

7.0

61.5

38,5

7.2

71.7

28,3

7.4

80.2

19,8

7.6

86,G

13,4

7.8

90.8

9,2

8.0

94.0

G.Q

Dilute die combined 1 v slack solutions toi liter with distilled htjí). [il [ Is calculated ¿nrrnnHin^ In rh.R t ÎFTifiRrrmn-E ]«üw»llw.Lrh Relation:

Dilute die combined 1 v slack solutions toi liter with distilled htjí). [il [ Is calculated ¿nrrnnHin^ In rh.R t ÎFTifiRrrmn-E ]«üw»llw.Lrh Relation:

pH = pJC +lo£ [{pro'.ûnaccipLûr)1 } prolan donor where pK = S^6 ai25T.

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