Abstract
Chronic myelogenous leukemia (CML) is the outcome of a chromosomal translocation and abl and bcr genes fusion. The present study is an attempt to implement a simple and inexpensive method for quantification of bcr-abl fusion transcripts in CML patients. Competitive RT-PCR technique was used to quantify bcr-abl transcripts. Internal control DNA piece was designed and synthesized from a non-human source, which was smaller than the target DNA in length but with the end identical to the main target piece. Following proliferation and purification, the transcriptions were quantified. To optimize reaction condition, competitive PCR was first performed separately on the target DNA with a given number of copies and a given number of copies of internal controls and the result of the reaction was electrophoresed for all concentrations of agarose electrophoreses gel. Results of simultaneous reactions of target RNA along with different number of copies of internal controls showed that the number of target RNA copies can e determined through comparing intensity of the color of DNA bands on the gel. The results showed that comparing with other methods like Real-time, the competitive RT-PCR is a relatively efficient and economic method to quantify bcr-abl transcripts.
Highlights
Chronic myelogenous leukemia is a clonal disorder of hematopoietic stem cell that leads to an increase in myeloid, erythroid, and megakaryocyte cell lines in peripheral blood and an increase of hyperplasia myeloid in bone marrow
Chronic myelogenous leukemia (CML) is associated with a cytogenetic disorder known as Philadelphia chromosome (Ph)
Two breakdowns occur at this region: one is after exon (e13) that results in el3a2 fusion (b2a2) and another is after exon (e14) that results in e14a2 (b3a2). Both of these fused mRNs are translated into a 210 kD protein (9210BCR-ABL) [3,4]. This type of fused protein is found in 30% of acute lymphoblastic leukemia (ALL)
Summary
Chronic myelogenous leukemia is a clonal disorder of hematopoietic stem cell that leads to an increase in myeloid, erythroid, and megakaryocyte cell lines in peripheral blood and an increase of hyperplasia myeloid in bone marrow. Two breakdowns occur at this region: one is after exon (e13) that results in el3a2 fusion (b2a2) and another is after exon (e14) that results in e14a2 (b3a2) Both of these fused mRNs are translated into a 210 kD protein (9210BCR-ABL) [3,4]. The 190kD protein is found in 50-70% of ALL patients as well Another rare breakpoint is (μ-BCR) that due to a breakdown at exon 19 (e19) leads to e19a2 fusion, which is translated into 23kD protein. This type of breakpoint is more common in chronic neutrophilic leukemia (CNL). This fusion is featured with increase in tyrosine kinase activity of bcr gene, which results in increase of cellular
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