![Figure 2. Molecular analysis of PRKAR1A-RARA fusion transcripts. (A) The diagram shows the exon structure of the human PRKAR1A and RARA mRNAs (GenBank Accessions NM_002734 and NM_000964, respectively), with translated regions indicated by wide rectangles. Exploded views of exons 2 and 3 of PRKAR1A, and exons 3 and 4 of RARA are indicated with the positions of PCR primers used, PRKAR1A-F1 (5′-gaaccatggagtctggc-3′), PRKAR1A-F2 (5′-ggttggagaaggaggag-3′), RARA-R34o1 (5′-ggcggaagaagcccttgcag-3′), and RARA-R3 (5′-cagccctcacaggcgctgac-3′). (B) PRKAR1A-RARA RT-PCR analysis using primers PRKAR1A-F1 and RARA-R34o1 resulted in the detection of 2 transcripts (PCR products of 444 bp [base pairs] and 344 bp) in the diagnostic bone marrow (Dx) of the index patient but no products were amplified from either the 11-month remission sample (11 mo), from Meg01 cell line (Meg), used as a negative control, or in a “no template” control (NTC). DNA molecular weight markers (M) were 100 base pair ladder (GE Healthcare, Buckinghamshire, United Kingdom). (C) Reamplification of the reactions shown in panel B with nested PCR primers PRKAR1A-F2 and RARA-R3 (that exclusively amplify fusions between PRKAR1A exon 3 and RARA exon 3) resulted in production of a single 256-bp product in the diagnostic sample. The absence of a product in the remission sample confirmed PRKAR1A-RARA molecular remission. (D) Partial sequence trace of the predominant in-frame PRKAR1A-RARA fusion transcript product showing the junction of the 2 genes as the result of RNA splicing from a cryptic splice donor within PRKAR1A exon 3. (E) Diagram showing the exon structures (narrow rectangles), open reading frames (wide rectangles), and partial amino acid sequence at the fusion junctions of the 2 main PRKAR1A-RARA transcripts present at diagnosis. PRKAR1A-derived exons and sequences are indicated by gray shading. The in-frame fusion (GenBank Accession EF428110) would be capable of encoding a 495–amino acid protein. The shorter frame-shifted fusion transcript (GenBank Accession EF428111) would encode a truncated RIα protein with 11 carboxy terminal amino acids (in italics) derived from the frame-shifted RARA exon 3.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/110/12/10.1182_blood-2007-06-095554/4/zh80240710050002.jpeg?Expires=1726815730&Signature=tSK2dJUDizrAPfxp8WmLHH4pvAznGz5GxqSufy9HLm9KkjaDL63OK4jSoO1XWnGVu3UQKHr~me293-x3jEWTovLLm6kQv~mhxeJCRQrWR3wVJjfKK8qMi6cXHIfxwg2SgRWRc5fiFeftsbfCijtViCk6MuctB9DF~WhiU8vsV7jMUKv58xzRuxtYiAJRtddr9ft67iAYkWtwyk0t2sllNuAzwAJKc8umlhCcrJB2~Dfu~HBDwVJ9VsrWNh0UEZs3ZVWLD-yBT-awvcsxjdjisoUzfMsDXUXjQRogvFWsK37XnY-AtvIDWzkAHLlOZPt1jNgMdTEBQfUUMK-MNABqZw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Molecular analysis of PRKAR1A-RARA fusion transcripts. (A) The diagram shows the exon structure of the human PRKAR1A and RARA mRNAs (GenBank Accessions NM_002734 and NM_000964, respectively), with translated regions indicated by wide rectangles. Exploded views of exons 2 and 3 of PRKAR1A, and exons 3 and 4 of RARA are indicated with the positions of PCR primers used, PRKAR1A-F1 (5′-gaaccatggagtctggc-3′), PRKAR1A-F2 (5′-ggttggagaaggaggag-3′), RARA-R34o1 (5′-ggcggaagaagcccttgcag-3′), and RARA-R3 (5′-cagccctcacaggcgctgac-3′). (B) PRKAR1A-RARA RT-PCR analysis using primers PRKAR1A-F1 and RARA-R34o1 resulted in the detection of 2 transcripts (PCR products of 444 bp [base pairs] and 344 bp) in the diagnostic bone marrow (Dx) of the index patient but no products were amplified from either the 11-month remission sample (11 mo), from Meg01 cell line (Meg), used as a negative control, or in a “no template” control (NTC). DNA molecular weight markers (M) were 100 base pair ladder (GE Healthcare, Buckinghamshire, United Kingdom). (C) Reamplification of the reactions shown in panel B with nested PCR primers PRKAR1A-F2 and RARA-R3 (that exclusively amplify fusions between PRKAR1A exon 3 and RARA exon 3) resulted in production of a single 256-bp product in the diagnostic sample. The absence of a product in the remission sample confirmed PRKAR1A-RARA molecular remission. (D) Partial sequence trace of the predominant in-frame PRKAR1A-RARA fusion transcript product showing the junction of the 2 genes as the result of RNA splicing from a cryptic splice donor within PRKAR1A exon 3. (E) Diagram showing the exon structures (narrow rectangles), open reading frames (wide rectangles), and partial amino acid sequence at the fusion junctions of the 2 main PRKAR1A-RARA transcripts present at diagnosis. PRKAR1A-derived exons and sequences are indicated by gray shading. The in-frame fusion (GenBank Accession EF428110) would be capable of encoding a 495–amino acid protein. The shorter frame-shifted fusion transcript (GenBank Accession EF428111) would encode a truncated RIα protein with 11 carboxy terminal amino acids (in italics) derived from the frame-shifted RARA exon 3.
