Part:BBa_K3577002
PCA3 mRNA
PCA3 is a kind of biomarker in prostate cancer tissue. The mRNA of PCA3 can be detected in the urine of patient with prostate cancer. PCA3 almost does not express in other cancers or just a little that can be ignored, so it has high specificity in prostate cancer. Also, the expression of PCA3 mRNA was significantly different between prostate cancer cells and normal cells, which shows a high sensitivity.
We searched PCA3 on the UCSC Genome Browser (http://genome.ucsc.edu/), a genome browser hosted by UCSC which offers access to genome sequence data. When determining which specific transcript of the gene to use, we chose the transcript with the most amount of exons. We then copied the sequence of one particular exon shared by multiple transcripts of the gene (NCBI Reference Sequence: NR_132312.1).
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal EcoRI site found at 139
- 12INCOMPATIBLE WITH RFC[12]Illegal EcoRI site found at 139
- 21INCOMPATIBLE WITH RFC[21]Illegal EcoRI site found at 139
- 23INCOMPATIBLE WITH RFC[23]Illegal EcoRI site found at 139
- 25INCOMPATIBLE WITH RFC[25]Illegal EcoRI site found at 139
- 1000COMPATIBLE WITH RFC[1000]
Usage and Biology
The PCA3 sequence we uploaded this time is only a fragment of the whole sequence, which is the region amplified by the PCA3 primers.
Results
We obtained three pairs of primers for PCA3, all pairs of primers of PCA3 were verified by PCR ( Polymerase Chain Reaction) before we start following research, the best one pair, PCA3-2,was used in further amplification (FIgure1).
The primer sequence was as follows:
PCA3-2F: GCAAGAGCCACAGAGGGAATG
PCA3-2R: GCGCACTCACCATGAAATGG
We added a T7 sequence at the 5’ end of forward primer to help start the process of transcription. For the reverse primer, we designed it with a trigger at the 5’ end, so as to add the trigger sequence to the end of PCR product during the process of PCR.
The sequences of these overhang-added primers are as follow :
T-PCA3-2F: taatacgactcactatagggGCAAGAGCCACAGAGGGAATG
T-PCA3-2R: gtttgaatgaattgtaggcttgttatagttatgtttGCGCACTCACCATGAAATGG
The results of PCR showed that the primers with overhang could also amplify PCA3 successfully without significantly affecting the amplification efficiency.
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