lose in primer viability and EDTA

Dr. Hiranya S. Roychowdhury hroychow at nmsu.edu
Thu Apr 5 20:35:38 EST 2001


Well, the EDTA in TE is at a low 1mM conc., which usually is taken into
consideration while doing PCR. I (and lot of others) use TE as the storage
buffer for DNA in every form. Keep in mind that, among those myriad of
manipulations we subject our Nucleic Acids to, it is not only the PCR
reaction that requires magnesium. 

Furthermore -- and I rhink most importantly -- every PCR reaction condition
should be worked out through a set of Mg++ concentrations, starting at zero
molar, which then makes the presence of mere 1mM EDTA immaterial.

At 09:04 AM 4/5/01 +0200, Christine Schneider wrote:
>I think the EDTA might cause a problem in the PCR reaction. EDTA chelates
(?) Mg,
>which is necessary for DNA-Polymerase.
>
>Christine
>
>
>"Dr. Hiranya S. Roychowdhury" schrieb:
>
>> Always reconstitute in a buffer. The concentration of the buffer may be as
>> loww as 1mM, but the NA's need to be buffered for longterm storage. 10mM
>> Tris.Cl, pH7.5 to pH8.0 is used almost universally. The TE may also be
used.
>>
>> At 11:15 AM 4/4/01 +0100, R. Jayakumar wrote:
>> >Hi.
>> >I have a strange problem with one of my set of primers.  This pair of
>> >primers is meant to amplify an antibiotic resistance gene.  the primers
work
>> >well only for two months or so, even though I use only a working stock
and I
>> >maintain the stock in -20C.  The primers somehow lose their viability
after
>> >2-3 months and I have to synthesize them again.  The same problem happens
>> >again after 2-3 months.  I reconstitute the lyophilized primers in
deionised
>> >water as soon as I receive it and store in -20C.  This is becoming a
>> >problem, because I have to synthesize them every so often.
>> >    can anybody suggest what could be the problem?
>> >sincerely
>> >jayakumar
>> >
>>
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>>
>||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
>> >||||
>> >R. JAYAKUMAR
>> >CSIR- Senior Research Fellow
>> >Dept. of Molecular Microbiology,
>> >School of Biotechnology,
>> >Madurai Kamaraj University,
>> >Madurai - 625 021.
>> >India
>> >email: jakku at linuxfan.com
>> >tel: +91-452-858471-374
>> >office fax(india)= +91-452-859105
>> >Efax(USA): (603)-688-4665
>>
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>>
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>> >||||
>> >"Life is a comedy for those who think and a tragedy for those who
>> >feel" --Emerson
>> >
>> >---
>> >
>> >
>> Dr. Hiranya Sankar Roychowdhury
>> College Asst. Prof.
>> Molecular Biology,
>> Dept. of Chemistry & Biochemistry
>> Box 30001 - 3MLS
>> New Mexico State University
>> Las Cruces, NM 88003
>>
>> Lab: (505) 646 4722
>> Office: (505) 646 8256
>> hroychow at nmsu.edu
>>
>> ---
>
>--
>______________________________________________________________
>
>Christine Schneider
>
>Ruhr-Universität Bochum
>AG Molekulare Genetik
>ND 06/756
>44780 Bochum
>
>Tel.: 0234 32-25629
>email: christine.schneider at ruhr-uni-bochum.de
>
>
>
Dr. Hiranya Sankar Roychowdhury
College Asst. Prof.
Molecular Biology,
Dept. of Chemistry & Biochemistry	
Box 30001 - 3MLS
New Mexico State University
Las Cruces, NM 88003

Lab: (505) 646 4722
Office: (505) 646 8256
hroychow at nmsu.edu

---




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