Genotyping: Help with calling tetranucleotide repeat peaks

Alfons Bardoel a.f.j.bardoel at med.uu.nl
Tue Mar 19 10:51:22 EST 2002


Hi,

We do find tetranucleotide repeats having alleles with 1 or 2 base
pair difference . So it's not always a +A addition. In the 3rd lane
of your example the peak is wider than in the 1st lane; small peak at
161 and high ones at 162 and 163. If you look in the 1st lane there
is a small peak at 161 and a high one at 162. In lane 1 I would
consider 162 as being the +A of 161;(so homozygote 162). In the 3rd
lane though trere's an extra peak and I would say this is a
heterozygote 162/163. You often see an extra peak 4 bp smalleer than
the "real" peak. You can see this in the 4th lane and in the 3rd lane
you can clearly see 2 small peaks (at 158 and 159), supporting this
being a heterozygote.
Because of the bad quality of the picture and the small number of
samples I wouldn't rely on it. Typing more samples and also taking
along CEPH controll samples gives more certainty.
Another thing is that you also have an allele at 164; so this makes
162, 163 and 164. I definitely would check your internal lanestandard
because this can ruin your sizecalling.

Good luck,

Alfons

>Hi Al'
>
>What do the genotypes look like in other individuals? this doesnt look
>like a nuclear family to me, if it was you could determine some of the
>genotypes from a haplotype. Why are your genotype bins so much wider for
>some alleles rather than others? As always... if in doubt they should the
>classed as unknown. Bad genotypes are squillion times worse than missing
>ones.
>
>Dont forget... repeats are mutable and as such might not appear truely
>Mendelian... at some point they have to change and will appear as bad
>inheritance. Also, I have seen  genotypes which are compounded by a rare
>ins/del polymorphism which give very strange but correctly inherited
>and very rare alleles.
>
>Hope some of this helps...
>
>Best wishes
>
>nick
>
>
>  On 15 Mar 2002, Alison Brown wrote:
>
>>  I'm having problems calling several of the allele peaks of a
>>  tetranucleotide repeat.  I have pasted a picture of several examples of
>>  it onto a web site in the hope that someone can help me with it.  Sorry
>>  about the quality of the picture - it was the best I could do.
>  >  http://dnaseq.bwh.harvard.edu/problem-tetra.html
>>
>>  The allele in question is 162 - if you can't read the labels that's the
>>  second peak of the first lane.
>>
>>  Lane 1 - looks good to me for both alleles ( peak sizes 150.08 and 162.00)
>>  Lane 2 - 2nd peak looks good at 166.93, but what should the peak at
>>  163.88 be called as?
>>  Lane 3 - Main peak is at 161.98 and 2nd at 162.97
>>  Lane 4 -  Peak is at 162.91
>>
>>  So is this +A addition?  Does this happen to tetra's, I thought it
>>  happened only to dinucleotide repeats.  Reverse primer has 7 base tail
>>  to force +A addition.
>>
>>  Any help is great!
>>  Thanks,
>>  Alison
>>
>>  --
>>  Alison Brown
>>  Laboratory Manager and Technical Director
>>  Harvard Partners Center for Genetics and Genomics
>>  High Throughput Genotyping Facility
>>  Brigham and Women's Hospital
>>  221 Longwood Avenue, LM114
>>  Boston, MA 02115.
>>
>>  Lab Tel: 617 732 5949
>>  Office Tel: 617 732 5561
>>  Fax: 617 264 5135
>>
>>
>>  ---
>>
>>
>
>end
>**************************************************
>Dr N I Leaves
>Mouse Sequencing
>MRC HGMP Resource Centre
>Hinxton
>Cambridge CB10 1SB
>tel: 01223 494557 (office) or 01223 494541 (lab)
>email: nleaves at hgmp.mrc.ac.uk
>**************************************************


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