Repository NIGMS Human Genetic Cell Repository
Subcollection Heritable Diseases
Lysosomal Storage Diseases
Protocols Protocol PDF
Cell Type Stem cell
Tissue Type Induced pluripotent stem cell
Transformant Reprogrammed (Sendai)
Race Caucasian
Family Member 1
Relation to Proband proband
Confirmation Molecular characterization after cell line submission to CCR
ISCN 46,XX[23].arr[hg19](1-22,X)x2
Species Homo sapiens
Common Name Human
Remarks Seizures; cortically blind; developmental delay; poor feeding; onset at age 6 months; deficient WBC lactosylceramide B-galactosidase activity; donor subject is homozygous for a 30 kb deletion beginning near the middle of intron 10 in the GALC gene resulting in the loss of the sequences encoded by exons 11-17 plus an additional 9 kb; same subject as GM06805 (lymphoid); induced pluripotent stem cell line derived from GM06806 (fibroblast).
Passage Frozen 15
 
Induced Pluripotent Stem Cell The frozen cell line submitted to the Repository was recovered and expanded. The expanded line was evaluated for viability surface antigen expression and alkaline phosphatase activity. Pluripotency was assessed via embryoid body (EB) formation. Steady-state mRNA expression patterns of undifferentiated iPSC and EBs were determined via real-time PCR. Characterization data are included in the Certificate of Analysis.
 
Gene GALC
Chromosomal Location 14q24.3-q32.1
Allelic Variant 1 606890.0002; KRABBE DISEASE, INFANTILE
Identified Mutation 30-KB DEL, IVS10; Rafi et al. (1995) stated that although most patients with globoid cell leukodystrophy have the severe infantile form, patients up to 50 years of age have been diagnosed in their laboratory. They reported that a large deletion, together with a polymorphic C-to-T transition at position 502 of the cDNA (counting from the A of the initiation codon), was responsible for a large number of disease-causing alleles in patients with Krabbe disease. Of 48 patients evaluated, 10 were found to be homozygous for the 502/del allele, 5 were heterozygous for this allele, 21 were heterozygous for the 502 mutation (presence of the deletion could not be confirmed), and 1 infantile patient was homozygous for the 502 mutation but at least 1 allele was not deleted. No patient was found to have the deletion without the 502 mutation. The deletion of the entire coding region located beyond exon 10 was first detected by failure to PCR amplify any cDNA sequences from that part of the gene. Intron 10 is the largest intron in the GALC gene and is estimated to be about 12 kb in length. In the patients with deletions, PCR amplification was successful for detecting only about 5 kb into intron 10. The large deletions starting within intron 10 result in the loss of coding information from 7 exons which represent about 15% of the 50-kD subunit and all of the 30-kD subunit. Normally, these 2 subunits are produced from the 80-kD precursor protein (Chen and Wenger, 1993). The occurrence of the deletion on the 502 allele and not on the normal allele indicates that the deletion event may have happened only once. The large deletion occurred particularly in patients with infantile Krabbe disease who have northern European ancestry; however, 1 homozygous patient had a Hispanic surname. Since more people have the 502 mutation and are carriers of classic Krabbe disease, it must be a polymorphism. However, the change from arginine to cysteine could result in improper folding of the GALC subunits due to abnormal disulfide bond formation. In a study of 64 unrelated patients with infantile Krabbe disease of Dutch (n = 41) or other European (n = 23) origin, Kleijer et al. (1997) found the large 30-kb deletion starting in intron 10 in 52% of the 82 GALC Dutch alleles and in 35% of the non-Dutch alleles. The 502T polymorphism, which had an allele frequency of 5.3% in a Dutch control panel, occurred in 65% of the GLD alleles. Analysis of patients and both parents in 26 of the Dutch families showed that the 30-kb deletion was invariably associated with 502T. However, 502T was also present on 40% of the GLD alleles with an as yet unidentified mutation, which is 7.5 times higher than its frequency in controls. This suggested that in addition to the 30-kb deletion, at least 1 other relatively frequent mutation had arisen on the the 502T GALC allele. See also 245200.0007.
 
Gene GALC
Chromosomal Location 14q24.3-q32.1
Allelic Variant 2 606890.0002; KRABBE DISEASE, INFANTILE
Identified Mutation 30-KB DEL, IVS10; Rafi et al. (1995) stated that although most patients with globoid cell leukodystrophy have the severe infantile form, patients up to 50 years of age have been diagnosed in their laboratory. They reported that a large deletion, together with a polymorphic C-to-T transition at position 502 of the cDNA (counting from the A of the initiation codon), was responsible for a large number of disease-causing alleles in patients with Krabbe disease. Of 48 patients evaluated, 10 were found to be homozygous for the 502/del allele, 5 were heterozygous for this allele, 21 were heterozygous for the 502 mutation (presence of the deletion could not be confirmed), and 1 infantile patient was homozygous for the 502 mutation but at least 1 allele was not deleted. No patient was found to have the deletion without the 502 mutation. The deletion of the entire coding region located beyond exon 10 was first detected by failure to PCR amplify any cDNA sequences from that part of the gene. Intron 10 is the largest intron in the GALC gene and is estimated to be about 12 kb in length. In the patients with deletions, PCR amplification was successful for detecting only about 5 kb into intron 10. The large deletions starting within intron 10 result in the loss of coding information from 7 exons which represent about 15% of the 50-kD subunit and all of the 30-kD subunit. Normally, these 2 subunits are produced from the 80-kD precursor protein (Chen and Wenger, 1993). The occurrence of the deletion on the 502 allele and not on the normal allele indicates that the deletion event may have happened only once. The large deletion occurred particularly in patients with infantile Krabbe disease who have northern European ancestry; however, 1 homozygous patient had a Hispanic surname. Since more people have the 502 mutation and are carriers of classic Krabbe disease, it must be a polymorphism. However, the change from arginine to cysteine could result in improper folding of the GALC subunits due to abnormal disulfide bond formation. In a study of 64 unrelated patients with infantile Krabbe disease of Dutch (n = 41) or other European (n = 23) origin, Kleijer et al. (1997) found the large 30-kb deletion starting in intron 10 in 52% of the 82 GALC Dutch alleles and in 35% of the non-Dutch alleles. The 502T polymorphism, which had an allele frequency of 5.3% in a Dutch control panel, occurred in 65% of the GLD alleles. Analysis of patients and both parents in 26 of the Dutch families showed that the 30-kb deletion was invariably associated with 502T. However, 502T was also present on 40% of the GLD alleles with an as yet unidentified mutation, which is 7.5 times higher than its frequency in controls. This suggested that in addition to the 30-kb deletion, at least 1 other relatively frequent mutation had arisen on the the 502T GALC allele. See also 245200.0007.
Remark Seizures; cortically blind; developmental delay; poor feeding; onset at age 6 months; deficient WBC lactosylceramide B-galactosidase activity; donor subject is homozygous for a 30 kb deletion beginning near the middle of intron 10 in the GALC gene resulting in the loss of the sequences encoded by exons 11-17 plus an additional 9 kb; same subject as GM06805 (lymphoid); induced pluripotent stem cell line derived from GM06806 (fibroblast).
No data is available
No data is available
Passage Frozen 15
Split Ratio 1:4
Temperature 37 C
Percent CO2 5%
Percent O2 AMBIENT
Medium Ham's F12 Medium/Dulbecco Modified Eagles Medium, 1:1 mixture with 2mM L-glutamine or equivalent
Serum 20% Knock-out Serum Replacement Not inactivated
Substrate Gelatin + Feeder Layer
Supplement Basic Fibroblast Growth Factor 10 ng/ml