Separating polymorphic amplified fragments of the same size

Yves Bertheau bertheau at inapv.inapg.inra.fr
Sat Nov 27 10:16:59 EST 1993


Hello,

We are currently ampifying 1 to 3 fragments (depending on the bacterial
strain we are studying) of 420 bp which appear polymorphic after restriction
(the summation of the size corresponds to 1-3 fragments).

We would like to sequence these fragments from several strains in order to
develop ASO probes. The possibilities seem to be:

- cloning the fragments and trying to find the polymorphic fragments but it
seems very fastidious...

- to separate the fragments and then to sequence them but how to do ?
	
	* TGGE or DGGE but developing TGGE protocols may be time consuming
	and I would like to avoid Etidium bromide staining for the effects
	of UV on DNA

	* chromatography ? ion exchange or size exclusion ? Which material
	is then available for such a work e.g. for HPLC?

	* other method(s)?

I would like to know your comments/advices about my problem.

Thanks in advance.

--
		Yves Bertheau
INRA INA P-G		Pathologie Vegetale	16 rue Claude Bernard	
75231 PARIS cedex 05	FRANCE	Tel +33 (1) 44.08.16.98 44.08.17.04
Fax: +33 (1) 44.08.17.00 Internet: bertheau at inapv.inapg.inra.fr



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