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P034 Polymorphism analysis of 24 short tandem repeats (STRs) in a large sample of Mexican mestizos from Mexico
78
P034
Abstracts / Human Immunology 78 (2017) 51–254
POLYMORPHISM ANALYSIS OF 24 SHORT TANDEM REPEATS (STRS) IN A LARGE SAMPLE OF MEXICAN MESTIZOS FROM MEXICO Hilario Flores-A a,b, Araceli Rodríguez a, Betsy A. González a,b, Andrea Munguia a, Aida Delgado b, Clara Gorodezky a,b. aDept of Immunology & Immunogenetics, InDRE, Secretary of Health, Mexico City, Mexico; b Fundación Comparte Vida, A.C., Mexico City, Mexico. Aim: Patterns of genetic variation within and between populations have provided novel insights into the origin and history of humans. STRs are versatile and informative markers that facilitated a great understanding of the diversity present in natural populations, being a common tool in forensics, paternity and anthropological studies. Data collection for such studies is conducted based on many groups. Mexican Mestizos are of mixed descent and are the result of admixture between the Indigenous people of Asian ancestry inhabiting the country, different Europeans, and to a lesser extent, Africans. We intended to describe the pattern of 24 STRs in Mexican Mestizos for the above purposes. Methods: STRs were typed in 817 Mestizo adults, born in Mexico, (55.9% males and 44.1% females), by using PowerPlex21, GenePrint 24 (Promega), Global filer and Identifiler (Thermofisher Scientific) systems. We investigated 24 autosomal STRs in genomic DNAs. The STRs were run in a 3500 Sequencer and the DNA fragment analysis was done with the Gene Mapper ID-X v1.4 software. Allele frequency(AF) was compared with other studies done for forensics. Linkage disequilibrium(D), hardy–Weinberg equilibrium (HWE), Power of Exclusion (PE), Power of Discrimination (PD) and R Pearson value were determined. Results: A total of 347 alleles at the 24 STR loci were found with corresponding AF ranging between 0.045 and 52.66. These STRs are highly polymorphic being the most diverse FGA, D18551, PentaE, D21511, SE33 and D6S1043 (31–18 alleles). The most frequent alleles >25% were; 10 at D2S441; 11 at D5S818; 11 at D7S820; 14 at D6S1043; 14 at D19S433; 19 & 20 at D12S391; 10 at D7S820. These were compared with data of Central Mexico and from 13 CODIS from Latin American and Caribbeans. Conclusions: The 24 autosomal STR loci provide highly informative polymorphic data, since the R Pearson value = 0.985 (SD = 0.02). Some alleles were in D, meaning they are not in random association in 13/24 STRs (D16S539, D12S391, D19S433, PentaD, FGA, TH1, TPOX, D18S51, PentaE, D21S11, D18S51, D3S1358, CSF1PO). The PD = up to 0.974, showing high heterozygosity and low probability of finding two individuals with the same genotypes and PE = 0.947 shows the power of detecting variation. STRs may also be used together, since many are in D, to solve deficient kinship cases or cases with mutations.
P035
EVALUATION OF A MARKER SET FOR CHIMERISM MONITORING BY NEXT-GENERATION SEQUENCING Loes A. van de Pasch, Bram Luiken, Erik H. Rozemuller, Maarten T. Penning. GenDx, Utrecht, Netherlands. Aim: Accurate monitoring of the chimeric status of patients after stem cell transplantation is essential for early detection of transplant rejection or relapse. Currently, micro-chimerism levels are mostly assessed by Short Tandem Repeat (STR) fragment length analysis on capillary electrophoresis systems. The major disadvantages of this technique are the poor, low-level sensitivity and the laborious data analysis. As more and more HLA-typing labs have access to NGS, it is efficient to also use NGS for chimerism monitoring. We hypothesize that NGS-based chimerism monitoring will have a higher sensitivity than conventional methods and have a wider dynamic range, also including the dynamic range of STR technology. Here, we present the preliminary results of chimerism testing by means of NGS and evaluate the use of different types of informative markers. Methods: PCR amplification primers have been designed for a set of polymorphic InDel and STR markers. Using these markers a number of artificial chimeric samples were amplified. NGS data was analysed with customized analysis tools designed to quantitate the absence, presence, and length of the markers. Results: Non-chimeric samples tested result in clear homo- or heterozygous patterns for all markers tested. No interfering noise levels were found higher than 0.05% of the total number of reads assigned to that amplicon. When conducting artificial chimerism tests in the range between 0.1% and 100%, all percentages included could clearly be differentiated. Correlation between allele ratio targeted and percentages found exceeded R2 = 0.995 for a number of markers. Conclusions: The results demonstrate that NGS-based chimerism monitoring is feasible, with a high sensitivity and wide dynamic range, for multiple markers tested.
L.A. van de Pasch: 5. Employee; Company/Organization; GenDx. B. Luiken: 5. Employee; Company/Organization; GenDx. E.H. Rozemuller: 5. Employee; Company/Organization; GenDx. 6. Stock Shareholder; Company/ Organization; GenDx. M.T. Penning: 5. Employee; Company/Organization; GenDx.
P034
Abstracts / Human Immunology 78 (2017) 51–254
POLYMORPHISM ANALYSIS OF 24 SHORT TANDEM REPEATS (STRS) IN A LARGE SAMPLE OF MEXICAN MESTIZOS FROM MEXICO Hilario Flores-A a,b, Araceli Rodríguez a, Betsy A. González a,b, Andrea Munguia a, Aida Delgado b, Clara Gorodezky a,b. aDept of Immunology & Immunogenetics, InDRE, Secretary of Health, Mexico City, Mexico; b Fundación Comparte Vida, A.C., Mexico City, Mexico. Aim: Patterns of genetic variation within and between populations have provided novel insights into the origin and history of humans. STRs are versatile and informative markers that facilitated a great understanding of the diversity present in natural populations, being a common tool in forensics, paternity and anthropological studies. Data collection for such studies is conducted based on many groups. Mexican Mestizos are of mixed descent and are the result of admixture between the Indigenous people of Asian ancestry inhabiting the country, different Europeans, and to a lesser extent, Africans. We intended to describe the pattern of 24 STRs in Mexican Mestizos for the above purposes. Methods: STRs were typed in 817 Mestizo adults, born in Mexico, (55.9% males and 44.1% females), by using PowerPlex21, GenePrint 24 (Promega), Global filer and Identifiler (Thermofisher Scientific) systems. We investigated 24 autosomal STRs in genomic DNAs. The STRs were run in a 3500 Sequencer and the DNA fragment analysis was done with the Gene Mapper ID-X v1.4 software. Allele frequency(AF) was compared with other studies done for forensics. Linkage disequilibrium(D), hardy–Weinberg equilibrium (HWE), Power of Exclusion (PE), Power of Discrimination (PD) and R Pearson value were determined. Results: A total of 347 alleles at the 24 STR loci were found with corresponding AF ranging between 0.045 and 52.66. These STRs are highly polymorphic being the most diverse FGA, D18551, PentaE, D21511, SE33 and D6S1043 (31–18 alleles). The most frequent alleles >25% were; 10 at D2S441; 11 at D5S818; 11 at D7S820; 14 at D6S1043; 14 at D19S433; 19 & 20 at D12S391; 10 at D7S820. These were compared with data of Central Mexico and from 13 CODIS from Latin American and Caribbeans. Conclusions: The 24 autosomal STR loci provide highly informative polymorphic data, since the R Pearson value = 0.985 (SD = 0.02). Some alleles were in D, meaning they are not in random association in 13/24 STRs (D16S539, D12S391, D19S433, PentaD, FGA, TH1, TPOX, D18S51, PentaE, D21S11, D18S51, D3S1358, CSF1PO). The PD = up to 0.974, showing high heterozygosity and low probability of finding two individuals with the same genotypes and PE = 0.947 shows the power of detecting variation. STRs may also be used together, since many are in D, to solve deficient kinship cases or cases with mutations.
P035
EVALUATION OF A MARKER SET FOR CHIMERISM MONITORING BY NEXT-GENERATION SEQUENCING Loes A. van de Pasch, Bram Luiken, Erik H. Rozemuller, Maarten T. Penning. GenDx, Utrecht, Netherlands. Aim: Accurate monitoring of the chimeric status of patients after stem cell transplantation is essential for early detection of transplant rejection or relapse. Currently, micro-chimerism levels are mostly assessed by Short Tandem Repeat (STR) fragment length analysis on capillary electrophoresis systems. The major disadvantages of this technique are the poor, low-level sensitivity and the laborious data analysis. As more and more HLA-typing labs have access to NGS, it is efficient to also use NGS for chimerism monitoring. We hypothesize that NGS-based chimerism monitoring will have a higher sensitivity than conventional methods and have a wider dynamic range, also including the dynamic range of STR technology. Here, we present the preliminary results of chimerism testing by means of NGS and evaluate the use of different types of informative markers. Methods: PCR amplification primers have been designed for a set of polymorphic InDel and STR markers. Using these markers a number of artificial chimeric samples were amplified. NGS data was analysed with customized analysis tools designed to quantitate the absence, presence, and length of the markers. Results: Non-chimeric samples tested result in clear homo- or heterozygous patterns for all markers tested. No interfering noise levels were found higher than 0.05% of the total number of reads assigned to that amplicon. When conducting artificial chimerism tests in the range between 0.1% and 100%, all percentages included could clearly be differentiated. Correlation between allele ratio targeted and percentages found exceeded R2 = 0.995 for a number of markers. Conclusions: The results demonstrate that NGS-based chimerism monitoring is feasible, with a high sensitivity and wide dynamic range, for multiple markers tested.
L.A. van de Pasch: 5. Employee; Company/Organization; GenDx. B. Luiken: 5. Employee; Company/Organization; GenDx. E.H. Rozemuller: 5. Employee; Company/Organization; GenDx. 6. Stock Shareholder; Company/ Organization; GenDx. M.T. Penning: 5. Employee; Company/Organization; GenDx.