Page 79 - IJPS-8-2

International Journal of
Population Studies β-thalassemia mutations in Himalayan population

was done for all 648 individuals. Molecular investigation comparison with the previous reports on β-thalassemia in
was done only for 64 individuals out of the total 648 different regions of India.
screened for HbA as their HbA levels were more than
2
2,
3.5%. The cutoff level of more than 3.5% of HbA is 4. Discussion
2
considered as the gold standard for β-thalassemia trait (Ou Numerous studies on thalassemia have been carried out
et al., 2011). All these 64 individuals were, thus, subjected throughout the globe. India, being a part of the thalassemic
to the detection of mutation through the amplification- belt, faces various problems, such as unawareness, lack
refractory mutation system-polymerase chain reaction of finance, resources, and social stigma for pre-marital
(ARMS-PCR), which is a simple method for detecting screening (Mohanty et al., 2013; Verma et al., 1997). Despite
any mutation involving single base changes or small the enormous amount of research and rapid developments
deletions. ARMS is based on the use of sequence-specific seen during the past decade, it still continues to be a national
PCR primers that allow an amplification of test DNA concern. According to the World Health Organization
only when the target allele is contained within the sample (WHO), the total thalassemic reported in India alone
(Little, 2001). Molecular analysis for the blood samples was accounts for 80 – 90% and is a common hemoglobinopathy
carried out at the Central Molecular Research Laboratory (Haritha et al., 2012). β-thalassemia is considered to be the
of SGRRIHMHS, Dehradun, India. Genomic DNA from cause of morbidity and mortality along with the source of
the blood samples was extracted using QIAamp DNA economic burden to the community (Piplani et al., 2013).
Blood Mini Kit (Qiagen, Valencia, CA, USA). As per the WHO (WHO, 2001) records, β-thalassemia

The amplification of the targeted sequences of the is a common hemoglobinopathy in India, and therefore,
extracted DNA was carried out through ARMS-PCR numerous studies have been carried out for Indian
technique. The five most common mutations prevalent population for the distribution of thalassemia mutation.
in India, that is, IVS 1-5 G-C, IVS 1-1 G-T, Codon 41/42 Since India represents an extremely heterogeneous
(-TCTT), Codon 8/9, and 619 bp deletion were considered population with numerous tribal pockets, diverse racial
for detecting mutations. The PCR reaction was carried origin, and high inbred diseases frequency among certain
out with initial denaturation at 94°C for 5 min, followed communities, the prevalence rate of thalassemia is very
by 30 cycles of denaturation at 94°C for 1 min, 1 min high for some particular communities. There are diverse
annealing at 60°C, extension at 72°C for further 1 min, and ethnicities residing in different parts of the Himalayan
final extension for 5 min at 72°C. After PCR reaction, the belt. A study carried out in western part of India, conveyed
products of PCR are obtained in the form of amplicons. To 22.7% thalassemia carrier women diagnosed with anemia
analyze the PCR products, 10 μl of amplicon was loaded (Mulchandani et al., 2008).
on 1.5% agarose gel along with 1 μL of 6X DNA loading Molecular confirmation was done to ascertain authentic
dye (ML015). In a separate well 3 μL of 50 bp DNA ladder reporting. In different local communities of India, the
(MBT084) was loaded as marker. The gel was then allowed reported range for the five most common β-thalassemia
to run for about 15 – 20 min at 125 volts and the results mutations was between 0.3% and 17% (Agarwal & Mehta,
were recorded in Gel Documentation System. 1982; Weatherall & Clegg, 2001a; 2001b; WHO, 2008).
The present research reported a frequency of 0.5% for
3. Results β-thalassemia mutations, which is within the range

The distribution of the five mutations, that is, IVS 1-5 G-C, described for the Indian population. The analysis of
IVS 1-1 G-T, Codon 41/42 (-TCTT), Codon 8/9, and 619 bp mutation spectrum revealed the highest prevalence for
deletion, among the 64 individuals was studied using ARMS- IVS1-5 (G-C) (18.75%) followed by Codon 8/9 (12.5%)
PCR. The most frequent mutation was IVS 1-5 (G-C), which and IVS 1-1 (G-T) (6.25%). The frequency of mutation
was present in 12 study subjects, accounting for 18.75% of was compared with the other reported mutations of Indian
the study subjects. While Codon 8/9 was found in 12.5% of population (Tables 1-4). Colah et al. (2009) discovered IVS
individuals, IVS 1-1 (G-T) was found to be present in 6.25% 1-5 (G-C) as a pre-dominant mutation throughout India,
of the study population. On the other hand, 619 bp deletion with 44.8% prevalence in the north and 71.4% in the east.
and Codon 41/42 (-TCTT) mutations were absent in our This pre-dominancy is in unison with our study where the
studied population. The allelic frequency for all the three frequency of mutation was found to be 18.75% for IVS 1-5
mutations, IVS 1-5 (G-C), IVS 1-1 (G-T), and Codon 8/9, (G-C).
was found to be same at 0.99. The distribution frequency In contrast to our study, a higher frequency (88.6%)
of the five investigated β-thalassemia mutations is shown of IVS I-5 (G-C) was reported in Orissa, 78.8% in
in Tables 1-4 according to the four major zones of India in Andhra Pradesh, and 77.2% in West Bengal (Colah

Volume 8 Issue 2 (2022) 73 https://doi.org/10.36922/ijps.v8i2.324