Abi 3500 gene analyzer

Manufactured by Thermo Fisher Scientific

Sourced in United States

The ABI 3500 Gene Analyzer is a capillary electrophoresis system designed for DNA sequencing and fragment analysis. It utilizes fluorescence-based detection technology to accurately identify and quantify DNA fragments.

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Lab products found in correlation

Aligner software, by CodonCode (9 mentions) Abi bigdye terminator kit, by Thermo Fisher Scientific (3 mentions) Hiseq 2500 sequencer, by Illumina (1 mentions) Nanoquant plate, by Tecan (1 mentions) Infinite m200 pro, by Tecan (1 mentions) Qiamp dna blood mini kit, by Qiagen (1 mentions) Qiaamp dna blood mini kit, by Qiagen (1 mentions)

4 protocols using abi 3500 gene analyzer

Genetic Analysis of Suspected CCDS

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Cr and GAA levels on DBS and the Cr/Crn ratio in the urine both were measured by LC-MS/MS (Acquity UHPLC-class Xevo TQD, Waters, USA) (8 (link),9 (link)). The peripheral blood of the patients with suspected CCDS and their parents were obtained for next-generation sequencing (NGS). Genomic DNA was obtained from venous (whole) blood of the index patients and their parents and sequenced using an Illumina HiSeq 2500 sequencer (Illumina, San Diego, USA). All detected mutations were confirmed by Sanger sequencing using ABI 3500 Gene Analyzer (Applied Bio-systems, USA).
Gene sequencing, variant interpretation, and reporting were performed according to the high-throughput sequencing and diagnostic protocol established by the Molecular Diagnosis Center at Pediatric Research institution (12 ). New variants were analyzed by Polyphen-2 (http://genetics.bwh.harvard.edu) and Mutation Taster (www.mutationtaster.org). A comprehensive assessment of each variant’s pathogenicity, including the inheritance pattern, was carried out according to the American College of Medical Genetics and Genomics criteria (13 (link)).

Sun W., Wang Y., Wu M., Wu H., Peng X., Shi Y., Xiao F., Wu B., Zhou W, & Lu W. (2023). Fourteen cases of cerebral creatine deficiency syndrome in children: a cohort study in China. Translational Pediatrics, 12(5), 927-937.

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Genetic Analysis of Blau Syndrome

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Case records of patients attending the Pediatric Rheumatology Clinic, Advanced Pediatrics Centre, Postgraduate Institute of Medical Education and Research, Chandigarh, India, were reviewed. A detailed analysis of the clinical profile, investigations, treatment, and outcome of children with BS was recorded. An ophthalmological examination of these children was carried out at the uveitis clinic, Advanced Eye Centre, PGIMER, Chandigarh. The genetic diagnosis was confirmed at Pediatric Immunology Laboratory, PGIMER, Chandigarh.
Peripheral blood samples were collected for molecular analysis after obtaining informed consent. Exon-4 of NOD2 gene was amplified using polymerase chain reaction (PCR) at controlled conditions using specific oligonucleotide primers, which were obtained from the Resource of Asian Primary Immunodeficiency Database (RAPID) (15 ). The PCR products were qualitatively examined by 1.5% agarose gel electrophoresis followed by purification and direct sequencing using the ABI Big Dye Terminator kit and ABI 3500 Gene Analyzer (Applied Biosystems, Foster City, CA, USA). Sequencing results were analysed using Codon Code Aligner software (Codon Code Corporation, Centerville, MA, USA).

Kumrah R., Pilania R.K., Menia N.K., Rawat A., Sharma J., Gupta A., Vignesh P., Jindal A.K., Rikhi R., Agarwal A., Gupta V., Singh S, & Suri D. (2022). Blau syndrome: Lessons learned in a tertiary care centre at Chandigarh, North India. Frontiers in Immunology, 13, 932919.

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Genetic Analysis of CASP3 SNP

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Two milliliters of peripheral blood was taken from cases and controls for genetic analysis. Deoxyribonucleic acid (DNA) extraction was done using QIAmp DNA Blood Mini Kit (QIAGEN, Hilden, Germany). DNA purity was checked using TECAN infinite M200 pro with Nanoquant plate (TECAN group, AG, Switzerland) taking the optical density of 260 nm and 280 nm. Samples were then stored at −80⁰C.
Genotyping for the SNP rs113420705 was done using polymerase chain reaction (PCR) and Sanger sequencing. PCR amplification reactions were carried out at controlled conditions as per the standard protocol with the help of an automated PCR thermal cycle (Applied Biosystems, Thermo Fisher Scientific, Massachusetts, USA). PCR cycle conditions are available on request.
PCR products were then subject to purification and direct sequencing using the ABI Big Dye Terminator kit and ABI 3500 Gene Analyzer (Applied Biosystems, CA, USA). Sequencing results were analyzed using Codon Code Aligner software (Codon Code Corporation, Centerville, (MA). Primer sequences used for the amplification of the CASP3 gene comprising the SNP of interest are illustrated in Table 1. Representative pictures of PCR and sequencing are depicted in Figure 1.

Das K.G., Bhattarai D., Kaur A., Kaur A., Kumrah R., Srivastava P., Rawat A, & Singh S. (2022). Association of single nucleotide polymorphism rs113420705 of CASP3 in children with Kawasaki disease from North India. Journal of Family Medicine and Primary Care, 11(9), 5404-5409.

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Genetic Analysis of CD40LG Gene

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Genomic DNA was extracted using QIAamp DNA Blood Mini Kit according to
the manufacturer’s protocol (Qiagen, Hilden, Germany) from EDTA
anticoagulated blood samples. All five exons and exon/intron junctions of the
CD40LG were individually amplified using polymerase chain reaction and specific
oligonucleotide primers which were obtained from Resource of Asian Primary
Immunodeficiency Database (RAPID). The PCR products were qualitatively examined
by 1.5% Agarose gel electrophoresis followed by purification and direct
sequencing using the ABI Big Dye Terminator kit and ABI 3500 Gene Analyzer
(Applied Biosystems). Sequencing results were analyzed using CodonCode Aligner
software (CodonCode Corporation, Centerville, MA).

Rawat A., Mathew B., Pandiarajan V., Jindal A., Sharma M., Suri D., Gupta A., Goel S., Karim A., Saikia B., Minz R.W., Imai K., Nonoyama S., Ohara O., Giliani S.C., Notarangelo L.D., Chan K.W., Lau Y.L, & Singh S. (2018). Clinical and molecular features of X-linked hyper IgM syndrome – An experience from North India. Clinical immunology (Orlando, Fla.), 195, 59-66.

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