Single Nucleotide Polymorphisms Of Olypolyglutamate Synthetase Gene In Hildren With Acute Leukemia

Objective： To detect the allelic frequencies and genotype distributionof coding single nucleotide polymorphisms (cSNPs) of folypolyglutamatesynthetase (FPGS) gene in Han population of children from Shenzhen cityof China, provide theoretical references for further study on detecting therelationship between FPGS genetic polymorphisms and methotrexate(MTX) sensitivity, and for personalized medicine in tumor chemotherapy.Methods： Bone marrow from91children with acute leukemia andperipheral blood from124children with simple upper respiratory tractinfection were collected. Total RNA was then extracted. The cDNA wasobtained by reverse transcriptase (RT)-polymerase chain reaction (PCR).The primers for first and nest PCR were designed by Premier Primer5.0and Oligo6.0. The cSNPs of FPGS gene were screened by PCR-denaturing gradient gel electrophoresis (DGGE) in all215samples. Allexperimental data were analyzed by SPSS13.0. The differences of allelicfrequencies between AL group and the normal control group and betweenthe two subtypes were tested by the chi-square test. Results： A novel missense mutation,502/490T>C (L151/101P), wasfound in the coding region of FPGS gene from the investigated children.And the cytosolic (L101P) and mitochondrial (L151P) variants displayedallelic frequencies of0.70%and0.47%in Han population of childrenfrom Shenzhen city of China, respectively. In the normal control group theallelic frequencies were1.21%(L101P) and0.81%(L151P). This mutationwas not detected in the AL children group. Four known cSNPs weredetected in our study,908/896C>T (L286/236L),1446/1434C>T(R466/416C),1513/1501G>A(S488/438N) and1516/1504C>T (A489/439V). The allelic frequencies were0.47%,0.70%,0.47%,0.23%ofmitochondrial variants, and0.47%,0,0,0.47%of cytosolic variants,respectively. There were no significant differences of allelic frequenciesbetween the AL children group and the normal control group, and betweenthe two variants via the chi-square test assessment. No association withsusceptibility to disease was observed. In addition, the study did not detectthe following27unknown cSNPs included in NCBI cSNPs database,253/241A>C,293281G>A,303/291C>T,335/323G>T,350/338C>T,380/368C>T,417/405C>T,426/414G>A,441/429C>T,471/459C>T,473/461C>G,756/744G>C,930/918A>G,962/950C>T,965/953C>T,1268/1256C>G,1360/1348T>A,1451/1439C>T,1466/1454G>A,1485/1473G>A,1535/1523T>A,1541/1529C>T,1544/1532C>T,1633/1621G>C,1643/1631A>G,1766/1754C>T,1799/1787C>T. Conclusion： PCR-DGGE combined with DNA sequencing is appliedto screen and detect FPGS gene cSNPs for the first time. DGGE is apractical technology that can be used in the detection of FPGS genemutation. A novel missense mutation,502/490T>C (L151/101P), is foundin the coding region of FPGS gene from the investigated children. And thecytosolic (L101P) and mitochondrial (L151P) variants display allelicfrequencies of0.70%and0.47%in Han population of children fromShenzhen city of China, respectively. Four known cSNPs are detected inour study,908/896C>T (L286/236L),1446/1434C>T (R466/416C),1513/1501G>A (S488/438N) and1516/1504C>T (A489/439V). The allelicfrequencies are0.47%,0.70%,0.47%,0.23%of mitochondrial variants,and0.47%,0,0,0.47%of cytosolic variants, respectively. The study hasnot detected the following27unknown cSNPs included in NCBI cSNPsdatabase,253/241A>C,293281G>A,303/291C>T,335/323G>T,350/338C>T,380/368C>T,417/405C>T,426/414G>A,441/429C>T,471/459C>T,473/461C>G,756/744G>C,930/918A>G,962/950C>T,965/953C>T,1268/1256C>G,1360/1348T>A,1451/1439C>T,1466/1454G>A,1485/1473G>A,1535/1523T>A,1541/1529C>T,1544/1532C>T,1633/1621G>C,1643/1631A>G,1766/1754C>T,1799/1787C>T.