Beta Hemoglobin DNA Sequencing

DNA sequencing of the coding regions, introns, and flanking sequences of the hemoglobin beta (HBB) gene in both directions to detect point mutations causing beta globin variants and beta thalassemia.

This test sequences bidirectionally a 2 kb contiguous region of the HBB (beta globin) gene from approximately bases 61941 to 63927 of Genbank U01317, corresponding to bases minus 195 to plus 1792 relative to the start of transcription. Patient sequences are compared against Genbank U01317 as genomic reference sequence to detect mutations. Mutations are reported at the genomic level, with base 1 corresponding to the start of transcription. Mutations that change an amino acid are reported at the protein level according to the sequence of the mature protein without its initiator methionine, and may also be described by the traditional name of the hemoglobin variant.

Correspondence between this mutation nomenclature system and the HGVS nomenclature system may be found at the web site http://globin.cse.psu.edu.

This test was developed and its performance characteristics determined by the Department of Laboratory Medicine at the University of Washington.

BLOOD:

• Adult: 5 mL LAVENDER TOP tube
• Child: 2 mL LAVENDER TOP tube
• Also acceptable: YELLOW TOP (ACD) tube
• Unacceptable: Heparin green top tubes

AMNIOCYTES or CULTURED CHORIONIC VILLUS CELLS:

• Two (2) T23 or One (1) T75 flask (minimum 1-T25 flask)

CHORIONIC VILLIS and/or TISSUE:

• In sterile tube or culture media, at least 5 mg tissue.

Prenatal testing requires concomitant testing for maternal cell contamination (see Online Test Guide, Lab Mnemonic Maternal Cell Contamination [MCC] for ordering and specimen requirements)

Blood: Refrigerate whole blood up to 1 week.

Amniocytes & cultured CVS cells: hold flasks at room temperature.

Chorionic villi &/or tissue: hold at room temperature.

Outside Laboratories: Ship whole blood at ambient temperature for receipt within 1 week of specimen collection.- For cultured amniocytes or chorionic villus cells and for CVS or other tissue, transport and store at room temperature within 24 hours of obtaining CV or removing cultured cells from incubator.