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High Quality Pre-made cDNA for PCR and RACE Applications
Jump-start
Gene Expression Analyses
• Analyze tissue-specific
gene expression
• Isolate the true 5’ and 3’ ends
of messages
• Identify tissue specific
splice variants
• cDNAs from various mouse
and human tissues available
FirstChoice® cDNAs significantly reduce the amount of time
and effort required for tissue-specific cloning and gene expression analyses.
FirstChoice cDNAs are available in RACE-Ready and PCR-Ready formats that were
developed specifically to simplify RACE and PCR applications.
Better Results With Longer cDNA
FirstChoice RACE- and PCR-ready cDNA products
begin with the same high quality FirstChoice Poly(A) RNA and
are a mixture of cDNA populations separately primed with random
decamers and oligo(dT). The RNA is converted into first strand
cDNA by an engineered M-MLV reverse transcriptase, ArrayScript (patent pending), that produces high yields of full-length cDNA
(Figure 1). The increased length of a mixed cDNA population should
be especially beneficial to researchers needing to identify the
5’ or 3’ ends
of messages, characterize genetic isoforms and splice variants,
or construct cDNA libraries. Both cDNAs are purified identically,
but the RACE-Ready cDNAs undergo additional pre-cDNA synthesis
processing to maximize representation of end sequences.
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Figure
1. Longer Average cDNA Length After
Reverse Transcription Using ArrayScript™. cDNAs
were prepared from 2–7 µg (after adapter
ligation; for RACE-Ready) or 5 µg (for PCR-Ready)
of FirstChoice® Poly(A) RNA. RNA was reverse transcribed
using either wild type M-MLV RT (200 U/µl) or ArrayScript
RT (200 U/µl), and oligo(dT) primers (A) or
random decamers (B). After column purification
to remove primers and small reaction products, cDNA length
was assessed using an Agilent® 2100 bioanalyzer.
The data represent the average cDNA length for 8 samples.
All samples showed an increase in length when reverse
transcribed with ArrayScript RT. Average length varied
with species, tissue, and RT primer used. Average cDNA
lengths were 31–93% longer when primed with oligo(dT) (A) and
7–15% longer when primed with random decamers (B).
As expected, the increase in average cDNA length is smaller
for random primed products, because primers that anneal
close to the termini can only produce finite-sized cDNAs.
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Application Ready cDNAs
FirstChoice PCR-Ready cDNAs contain sufficient cDNA for 50 PCR
reactions and are supplied with control primers. Both PCR-Ready
cDNA and RACE-Ready cDNA can be used for routine PCR, but PCR-Ready
cDNA is a more economical choice, with no compromise in performance. FirstChoice RACE-Ready cDNAs contain cDNA for 30 RLM-RACE reactions and include
adapter primers for both 5’ and 3’ end-specific amplification and
controls. Both FirstChoice RACE-Ready and PCR-Ready cDNAs are available from
a variety of human and mouse tissues.
Scientific Contributors
Dana Broussard, Bret Schneider • Ambion, Inc.
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