PaeI (SphI)

Kód produktu: ER0601 Kód výrobce: ER0601 Kód dodavatele: {5521CB4E-8D92-455D-BCF0-C7FEFC59F9CF} Výrobce: Life Technologies Czech Republic s.r.o.

`5'...G C A T G↓C...3' 3'...C↑G T A C G...5'`

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Detailní popis

Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.

Highlights

Superior quality – stringent quality control and industry leading manufacturing process
Convenient color-coded Five Buffer System
Includes universal Tango buffer for double-digestions
BSA premixed in reaction buffers
Wide selection of restriction endonuclease specificities

Applications

Molecular cloning
Restriction site mapping
Genotyping
Southern Blot
Restriction fragment length polymorphism (RFLP)
SNP

Note

For methylation sensitivity refer to product specifications.

Compatible Ends	Hin1II, XceI.
Conditions for 100% Activity	1X Buffer B: 10 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl₂, and 0.1 mg/mL BSA Incubate at 37°C
Digestion of Agarose-embedded DNA	Minimum 5 units of the enzyme are required for complete digestion of 1 µg of agarose-embedded lambda DNA in 16 hours.
Double Digestion	Perform double digestion using DoubleDigest.
Hazardous	No
Isoschizomers	Search for commercial isoschizomers using REsearch.
Storage Buffer	PaeI is supplied in: 10 mM potassium-phosphate (pH 7.0 at 25°C), 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 0.15% Triton X-100, 0.5 mg/mL BSA, and 50% (v/v) glycerol.
Storage Condition	-20 C

Reaction conditions

Recommended buffer for 100% activity	Optimal temperature	Enzyme activity in Thermo Scientific buffers, %						Tango buffer for double digestion
Recommended buffer for 100% activity	Optimal temperature	B (blue) 1X	G (green) 1X	O (orange) 1X	R (red) 1X	Tango (yellow) 1X / 2X		Tango buffer for double digestion
B (Blue)	37°C	100	50-100	0-20	0-20	50-100	0-20	1X

Methylation Effects

Dam: never overlaps - no effect.
Dcm: never overlaps - no effect.
CpG: may overlap - no effect.
EcoKI: never overlaps - no effect.
EcoBI: may overlap - blocked.

Methylation type	Sequence	Cleavage effect
CpG	5'...`m5C GCATGm5C G`...3' 3'... `Gm5CGTAC Gm5C`...5'	No effect
EcoBI (TGA(N)₈TGCT)	5'...`TGm6A(N)₅GCA TGCT`...3' 3'...`AC T(N)₅CGTm6ACGA`...5'	Blocked

Cleavage efficiency close to the termini of PCR fragments

bp from the recognition site to fragment end
1	2	3	4	5
0	0-20		20-50	50-100

Number of recognition sites in DNA molecules

Lambda	ΦX174	M13mp18/19	pBR322	puc18/19	pUC57
6	0	1	1	1	1
pTZ19R/U	pTZ57R	pBluescriptIIKS(±)	pBluescriptIISK(±)	pACYC177	pACYC184
1	1	0	0	0	1

New sites generated by ligation

Newly generated recognition sites resulting from ligation of protruding compatible DNA ends

Recognition Sequence	Second Enzyme	Enzymes recognizing newly generated recognition sequence
`GCATG^C`	Hin1II (NlaIII)/FastDigest NlaIII (Hin1II) (CATG)	CviAII FaiI FatI Hin1II (NlaIII)/FastDigest NlaIII (Hin1II)
	XceI (NspI)/FastDigest NspI (XceI)* (`RCATG^C`)	Cac8I CviAII FaiI FatI Hin1II (NlaIII)/FastDigest NlaIII (Hin1II) PaeI (SphI)/FastDigest SphI (PaeI) XceI (NspI)/FastDigest NspI (XceI)
	XceI (NspI)/FastDigest NspI (XceI)* (`RCATG^T`)	CviAII FaiI FatI Hin1II (NlaIII)/FastDigest NlaIII (Hin1II) XceI (NspI)/FastDigest NspI (XceI)

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5'...G C A T G↓C...3' 3'...C↑G T A C G...5'

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