Analysis Of Existing Screening Techniques And Screening Of New Marker Genes For Down Syndrome

Down syndrome(DS),known as Trisomy 21 or congenital as well,which is the earliest main chromosomal disease and one of the most common serious birth defects.The rate of the disease on neonatal morbidity is about 1.25‰ and there is no effective or active treatment method for the disorder at present.Neonates with DS may bring families as well as the society huge economic burden and mental pain,as a result,preventing the birth of neonates with DS clinically is the main strategy.Clinical measures in the prevention for DS are mainly performed by serological prenatal screening in the early or middle periods of pregnancy,and then confirmed with cytogenetic techniques such as amniotic fluid puncture and villus puncture.However,this strategy result has a problem that it is in a high screening rate but a low diagnostic rate.In this research the current common screening and diagnostic technologies were estimated to study their performance on adaptation and detection in the prevention of DS,and new screening methods at the level of transcriptome was used to find new screening factors,so that to provide theoretical basis for establishing more effective diagnostic tests in the future.I.Performance analysis of the common screening methods for DSIn this study,all the pregnant women in clinical examination were screened by serological tests,meanwhile the gravida who were high-risk of DS were detected with non-invasive prenatal testing(NIPT).And karyotype analysis of amniocentesis,as the diagnosis criterion,was applied in the test.Besides,fluorescence in-situ hybridization(FISH)and chromosomal microarray analysis(CMA)were also used as complementary methods.The results were shown as follows.1.Through the means of serological test on 41267 pregnant women,there only1621 ones were positive,which accounts for 3.93%.Finally,554 pregnant women(34.18%)were tested by amniocentesis test and 37 pregnant women were diagnosed with DS,the diagnosis rate was 6.68%.Serological test was simpler and more common with the highest rate of acceptance(94.09%)among pregnant women but a lower rate of screening effectiveness.its positive predictive value was only 1.32%.2.Through the means of the tested with NIPT on 2667 pregnant women,19 of them were positive and the positive rate was 7.12%.The positive pregnant women were further tested by amniocentesis test,of which 18 patients were diagnosed with DS,the diagnosis rate was 94.73%,which was increased by 14.18 times.NIPT screening technology was very difficult and experiment be used as a common screening method,and its informed consent rate was 21.97%.But the positive predictive value of NIPT was as high as 94.74%.3.Total 237 specimens were analyzed with amniocentesis culture technique as the golden standard,3 of them were failed and the rate of failure was 1.27%.Besides,one case with partial deletion DS was missing.This technique needed an averageincubation period as long as 21 days,and was also difficult to be mastered and molecular detection technology was required to supplement.4.Statistics of 237 samples detected simultaneously by FISH technology and amniocentesis technology showed that the detection success rate of FISH technology was 100% with a detection cycle of 24-48 hours.Unlike amniocentesis with longer term of culture and higher failure rate,FISH was more rapid,but the defection was disability on detecting of chimera and limited application extent.5.CMA technology was showed with higher resolution rate and sensitivity after60 samples of amniotic fluid are detected by CMA and amniocentesis simultaneously.Detected 7 cases of trisomy 21,including 1 case of trisomy 21 that was missed by cytogenetics technology,and the diagnosis rate increased by 1.66%.However,CMA was difficult to diagnose and interpret on chromosome micro-deletion and micro-repetition with unclear clinical information,therefore only could be used as a complementary test.?.Transcriptomics study on the blood and placenta sample of pregnant women with DS1.The results of above research indicated the current various screening and diagnostic methods could be mutually complement,but the problem of clinical diagnosis with low diagnostic rate was still not to be completely solved.With the rapid development of genome sequencing technology,it is believed that genome sequencing can be used as a new diagnostic approach.In this research,the samples of blood and placenta from four groups of pregnant women with DS fetus in older age,DS fetus in younger age,normal fetus in older age and normal fetus in younger age were collected for transcriptome screening respectively,the genes related to Down's syndrome at the transcriptome level was screeninged.1.Total 30.07 Gb of gene data was obtained in the transcriptome sequencing and the proportion of Clean Reads was 98.15%,and had sequencing has reached the standard as well.The differential expression of genes in each group was apparent through result analysis.In the analysis of blood sample,the number of differentially expressed genes between the group of older pregnant women with DS fetus and the group of older pregnant women with normal fetus was greater than that between the group of younger pregnant women with DS fetus and the group of younger pregnant women with normal fetus,while in the analysis of placenta sample,the number of differentially expressed genes between the group of younger pregnant women with DS fetus and the group of younger pregnant women with normal fetus was greater than that between the group of older pregnant women with DS fetus and the group of older pregnant women with normal fetus.Up-regulated genes were more common in groups of women of older age.Besides,there were more differentially expressed genes in placenta tissues between the elder and the younger with DS fetus groups than those in the blood.2.All the identified differentially expressed genes were performed with GO function analysis.In the comparison analysis from the blood and placenta sample of groups of pregnant women with DS or normal fetus,3987 GO terms were found 3102 terms in biological processes,336 terms in cellular components,549 terms inmolecular functions.There are 138 terms identified within the comparison of four groups: 87 terms inbiological processes,35 terms in cellular components,and 16 terms in molecular functions.3.Total 104 pathways were enriched within the four groups by the KEGG classification.Compared with pregnant women with DS and normal fetus,3pathways were identified significantly,they were complement and coagulation cascades,Malaria and ECM-receptor interaction.Four pathways were found in blood and placenta samples from elderly groups and they were nicotine addiction,cAMP signaling pathway,adrenergic signaling in cardiomyocytes,aldosterone synthesis and secretion.Two pathways were enriched of the younger age group that were autoimmune thyroid disease and arachidonic acid metabolism.4.Four genes associated with DS in blood and placenta samples of older group were identified and they were CRIP2,NR4A1,PFKFB3 and FAM118 A.HLA-DRB5 and GNLY were found to be associated with DS in blood and placenta samples of younger groups.In the blood sample from older and younger groups with DS FCGR3 B and HLA-DRB4 were identified.With the confirmation of Q-PCR test,the expression levels of those genes were consistent with the results by transcriptome sequencing.5.The above 8 genes were verified with the clinical analysis,CRIP2,NR4A1,HLA-DRB5 and HLA-DRB4 could be used as candidate genes,which provideed potoential markers for non-invasive examination of DS.