Subtractive PCR?

murianap muriana at
Thu Dec 19 11:49:11 EST 1991

In article <1991Dec18.224447.8529 at>, rcbrown at (Robert Brown) writes:
>In article <1991Dec18.213347.27747 at> cbeall at (Clifford J Beall) writes:
>>Dear Methods people,
>>Does anyone have a method for subtracting cDNA prior to PCR?
>>We are using degenerate primers and want to clone a cell specific
>>message.  It seems one could hybridize with [+] cDNA with [-] RNA,
>>remove hybrids, then treat the remainder with RNAse.  Is HAP still the
>>method of choice for hybrid removal and does anyone know conditions
>>for this?  Thanks in advance.
>>				Cliff Beall
>>				Ohio State Biotech. Ctr.
>Invitrogen sells a kit for subtractive hybridization using biotinylation of
>the [-] RNA and extraction of the hybrid into phenol after binding

Here are some references regarding cDNA subtraction:

Nucleic Acid Res, 14:10027, 1986 ; Welcher, Torres, & Ward
	"Selective enrichment of specific DNA, cDNA, and RNA 
	sequences using biotinylated probes, avidin, and 
	copper-chelate agarose.

Nucleic Acids Res, 17:807, 1989;  Kuze, Shimizu, and Honjo
	"A new vector and RNase H method for the subtractive hybridization"

Nucleic Acids Res, 18:4833, 1990;  Rubenstein et al.
	"Subtractive hybridization system using single-stranded phagemids
	with directional inserts"

Anal. Biochem, 187:364-373, 1990; Fargnoli, Holbrook, and Fornace
	"Low-ratio hybridization subtraction"

PNAS, 87:2720-2724, 1990; Wieland, Bolger, Asouline and  Wigler
	"A method for difference cloning: gene amplification following 
	subtractive hybridization"

Anal. Biochem., 186:369-373, 1990; Barr and Emanuel
	"Application of a subtraction hybridization technique involving 
	photoactivatable biotin and organic extraction to solution
	hybridization analysis of genomic DNA"

Nucleic Acids Res, 19:1954, 1991; Swaroop et al.
	"A simple and efficient cDNA library subtraction procedure: 
	isolation of human retina-specific cDNA clones"

Hope this helps,
Peter Muriana

    Name: murianap
Internet: muriana at
   Phone: 317-494-8284

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