Analysis of β/α Globin Ratio by Using Relative qRT‐PCR for Diagnosis of Beta‐Thalassemia Carriers

Ranjbaran, Reza; Okhovat, Mohammad Ali; Mobarhanfard, Arash; Aboualizadeh, Farzaneh; Abbasi, Mozhdeh; Moezzi, Leili; Golafshan, Habib Allah; Behzad-Behbahani, Abbas; Bagheri, Mansooreh; Sharifzadeh, Sedigheh

doi:10.1002/jcla.21594

Cited by 9 publications

(12 citation statements)

References 13 publications

(23 reference statements)

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“…Onestep real-time RT-PCR for quantification of α-and β-globin mRNA was performed using SensiFast™ SYBR No-ROX One-Step Kit (Bioline USA Inc., Taunton, MA, USA) on Eco™ Real-Time PCR System (Illumina Inc., CA, USA) with α-globin mRNA specific primers; αG36 (5′-CCGACAAG ACCAACGTCAAGG-3′) and αG62 (5′-TCGAAGTGCGGGAAGTAGGTC-3′) and β-globin mRNA specific primers; G52 (5′-CCTGAGGAGAAGTCTGCC GTTAC-3′) and G82 (5′-AAAGGTGCCCTTGAGG-TTGT-3′). The relative quantification of α-and β-globin mRNA was normalized by normal α-and β-globin mRNA control and calculated by (efficiency) −ΔΔCT method as described previously [11,12]. The α/β globin mRNA ratios were determined on representative samples of normal, Hb E trait, β + -thalassemia trait (NT-28; A-G), Hb Dhonburi trait and β 0 -thalassemia trait (codons 41/42, −TCTT).…”

Section: Rna Analysismentioning

confidence: 99%

A large cohort of β+-thalassemia in Thailand: Molecular, hematological and diagnostic considerations

Yamsri

Singha

Prajantasen

et al. 2015

Blood Cells, Molecules, and Diseases

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Section: Rna Analysismentioning

confidence: 99%

A large cohort of β+-thalassemia in Thailand: Molecular, hematological and diagnostic considerations

Yamsri

Singha

Prajantasen

et al. 2015

Blood Cells, Molecules, and Diseases

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“…Others approaches using quantitative real-time PCR have showed a significant differences of the HBA/HBB mRNA globin ratio in reticulocytes between control and b-thal groups (Ranjbaran et al, 2013). The direct measurements of a/non a-globin biosynthetic or HBA/non HBA mRNA ratios are difficult to obtain routinely and they also only reflect the reticulocyte stage.…”

mentioning

confidence: 99%

Red blood cells free α‐haemoglobin pool: a biomarker to monitor the β‐thalassemia intermedia variability. The ALPHAPOOL study

et al. 2017

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The severity of β-thalassaemia (β-thal) intermedia is mainly correlated to the degree of imbalanced α/non α-globin chain synthesis. The phenotypic diversity of β-thal depends on this imbalance and reflects all possible combinations of α- and β-globin genotypes, levels of fetal haemoglobin (HbF) and co-inheritance of other modulating factors. This study aimed to demonstrate the validity of a new surrogate of α/non α-globin biosynthetic ratio by measuring the soluble α-Hb pool in lysed red blood cells. Our results confirm that the α-Hb pool measurement allows a good discrimination between β-thal intermedia patients, controls and α-thal patients (P < 0·003). Receiver operator characteristic analyses revealed an area under the curve of 0·978 for the α-Hb pool measurement at a threshold of 120 ng free α-Hb/mg of total Hb/ml of haemolysate (ppm) with a sensitivity and specificity of 86% and 100%, respectively, to discriminate between β-thal and not β-thal subjects. Significant correlations were observed between the α-Hb pool and biological parameters of β-thal, the most significant association being observed with red cell hexokinase activity. This study indicates that the α-Hb pool could be a new marker for assistance in diagnostic orientation of β-thal intermedia patients and may be clinically useful for monitoring the evolution of the disequilibrium of globin synthesis in response to treatments.

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“…The biological importance of this process is illustrated by common, natural occurring mutations that shorten the t 1/2 value of human α-globin mRNA, effecting a 5-fold reduction in its accumulation within 3 days of transcriptional silencing (Morales et al, 1997; Weiss and Liebhaber, 1994). Similar regulatory defects that reduce the level of β-globin mRNA would be predicted to lead to a significant discordance in α- and β-globin chain synthesis that defines congenital genetic conditions that include the β thalassemias (Chaisue et al, 2007; Ranjbaran et al, 2013). …”

Section: Discussionmentioning

confidence: 99%

AUF-1 and YB-1 independently regulate β-globin mRNA in developing erythroid cells through interactions with poly(A)-binding protein

Zalen

Lombardi

Jeschke

et al. 2015

Mechanisms of Development

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The normal expression of β-globin protein in mature erythrocytes is critically dependent on post-transcriptional events in erythroid progenitors that ensure the high stability of β-globin mRNA. Previous work has revealed that these regulatory processes require AUF-1 and YB-1, two RNA-binding proteins that assemble an mRNP β-complex on the β-globin 3′UTR. Here, we demonstrate that the β-complex organizes during the erythropoietic interval when both β-globin mRNA and protein accumulate rapidly, implicating the importance of this regulatory mRNP to normal erythroid differentiation. Subsequent functional analyses link β-complex assembly to the half-life of β-globin mRNA in vivo, providing a mechanistic basis for this regulatory activity. AUF-1 and YB-1 appear to serve a redundant post-transcriptional function, as both β-complex assembly and β-globin mRNA levels are reduced by coordinate depletion of the two factors, and can be restored by independent rescue with either factor alone. Additional studies demonstrate that the β-complex assembles more efficiently on polyadenylated transcripts, implicating a model in which the β-complex enhances the binding of PABPC1 to the poly(A) tail, inhibiting mRNA deadenylation and consequently effecting the high half-life of β-globin transcripts in erythroid progenitors. These data specify a post-transcriptional mechanism through which AUF1 and YB1 contribute to the normal development of erythropoietic cells, as well as to non-hematopoietic tissues in which AUF1-and YB1-based regulatory mRNPs have been observed to assemble on heterologous mRNAs.

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Analysis of β/α Globin Ratio by Using Relative qRT‐PCR for Diagnosis of Beta‐Thalassemia Carriers

Cited by 9 publications

References 13 publications

A large cohort of β+-thalassemia in Thailand: Molecular, hematological and diagnostic considerations

A large cohort of β+-thalassemia in Thailand: Molecular, hematological and diagnostic considerations

Red blood cells free α‐haemoglobin pool: a biomarker to monitor the β‐thalassemia intermedia variability. The ALPHAPOOL study

AUF-1 and YB-1 independently regulate β-globin mRNA in developing erythroid cells through interactions with poly(A)-binding protein

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