6 replies to this topic

[cbraval]

I have been having a lot of trouble tracking down sequence information for bovine AQP-1 for RT-Q-PCR.  Anyone have a suggestion to track it down.  Pubmed and google did not help my cause even with vague string searches.

[pcrman]

Why don't you design your own? the mRNA sequence for that gene is available from databases.

[cbraval]

Well from the people in my department it seemed prudent to get a published sequence to ensure it works on my cells.  However if I do need to design my own what is the general success of such attempts?  Will I have a lot of troubleshooting or does it work pretty straightforward?

[BryanC]

ABI has "made to order" Taqman probes for bovine aquaporin1

[cbraval]

https://products.app...EKeywordResults

I found AQP-1 from ABI for bovine species.  Why does it reference colton blood group?  Also when it says made to order what does that mean still needs to be specified?  I am sorry for the naive questions I am doing this for the first time and just want to be clear.  Thanks for your time and responses.

[Trof]

AQP1 is the gene coding the antigen in Colton blood group system. It's just the gene identification (but it tells you there are multiple variants of the gene).
Made to order means that assay is manufactured at the time of order, not just ready to ship, they differ in delivery time.
You only need to specify probe label and scale.

By the way I noticed you selected an assay with _g on the end, that means it detects gDNA. You usually don't want that for mRNA quantification, choose one of the assays with _m on the end, those have probe on the exon-exon boundary.

[BryanC]

What Trof said.


And to expand more....

Quote

Assay nomenclature: The first two letters in the
Assay ID represent the species. The letters after
the underscore give information about the assay
placement.

Example Assay ID: Hs00380483_m1 (Human assay
#00380483, which spans an exon junction).

Hs – Homo sapiens
Mm – Mus musculus
Rn – Rattus norvegicus
Dm – Drosophila melanogaster
At – Arabidopsis thaliana
Ce – Caenorhabditis elegans
Cf – Canis familiaris
Rh – Macaca mulatta (Rhesus)
Dr – Danio rerio (Zebrafish)


The gene expression Assay ID suffix indicates the
assay placement:

• “_m” indicates an assay whose probe spans an
exon junction and will not detect genomic DNA.

• “_s” indicates an assay whose probes and primers
are designed within a single exon. Such assays will,
by definition, detect genomic DNA.

• “_g” indicates an assay that may detect genomic
DNA. The assay probe and primers may also be
within a single exon.

• “_mH”, “_sH”, or “_gH” indicates that the assay
was designed to a transcript belonging to a gene
family with high sequence homology. The assays
have been designed to give between 1,000–
30,000-fold greater discrimination from the
closest homologous transcript.

• “_u” indicates an assay whose amplicon spans an
exon junction and the probe sits completely in one
of the spanned exons.

• “_ft” indicates the assay is designed to detect
fusion transcripts that result from chromosomal
translocation. The probe and one primer are on one
side of the fusion transcript breakpoint; the second
primer is on the other side of the fusion transcript
breakpoint. The assay does not detect genomic DNA.

• “_at” indicates the assay is designed to detect
a specific synthetic RNA transcript with a unique
sequence that lacks homology to current annotated
biological sequences.

http://www3.appliedb.../cms_050267.pdf

Other support info: https://products.app...;tab=Literature