Poor capillary transfer?

Wasun Chantratita asmsi002 at CMU.CHIANGMAI.AC.TH
Tue Mar 22 09:50:25 EST 1994


 

On 18 Mar 1994, Tony Sanchez wrote:

> 		Until we get our vacuum blotter, we are transfering our 
northerns
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Please try "Downward Capillary Transfer". We now follow the protocol from 
Ming Yi Zhou and his colleague, Biotecniques, vol.16, No. 1 (1994). Using 
cellulose sponge soaked with 10XSSC, you can have your high molecular 
weight DNA (>12Kb)transfered from agarose gel to nylon membrane within 2 hrs.

Forget about buying vacuum blotter. Only 10 Bath (25 Bath = $1) I can 
buy a big piece of cellulose sponge in Thailand. Cut it to fit the size of 
your gel, the rest you can use in the Lab for claning you glassware. The 
tickness of the sponge should be 3-4 cm. My fovorite size is 
12X12X5 cm3. At this size could absorb 200 ml 10X SSC. It is also 
autoclavable.
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> using 10xSSC capillary transfer overnite. But what we are finding is that
> even after 16 hr transfer, only the 18S band is transfering, while the
> higher stuff is staying very nicely in the gel.

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We usually spend 3 hr for transfer. Though, we have not yet tried with RNA 
but Dr. Zhou showed the nice picture of transfering 28s and 18s of mRNA 
of Rat.
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 I have made sure that
> there are enough paper towels and enough buffer in the reservoir. I was
> thinking that maybe the weight I put on top of the paper towels is too
> heavy, thus crushing the gel and preventing transfer of HMW bands. Any
> other ideas?
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There is no weighted wet paper towels, no gel flattening that might 
retard the transfer process.

Wasun

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Wasun Chantratita, Ph.D.                        Phone:053-221122 Ext 5086,5068
Department of Clinical Microbiology             Fax:  053-221890 
Faculty of Associated Medical Sciences       Email:asmsi002 at cmu.chiangmai.ac.th
Chiang Mai University
Chiang Mai 50200
Thailand
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