[Optimizing of cDNA preparation for next generation sequencing].

Next generation sequencing has already been used for genomic analysis of microorganis, human being, animals, and plants. Sample preparation is prerequisite and most important for large-scale sequencing. There are two major interferences for large-scale sequencing, polyA and abundant genes' concealment for rare genes. In order to solve these problems, we used total RNA extracted from violaceae leaves to produce double stranded cDNA. DSN nuclease was used to treat the ds cDNA prior to removing the polyA. Randomly sequencing 100 clones of the treated cDNA showed that there were 94 independent clones in the treated sample, and the sequences did not contained polyA. However, only 62 independent clones were found in the untreated sample, and 15 of the sequencing files were affected by polyA. By randomly sequencing of the treated cDNA, we also found two clones encoded two interested genes. We failed to isolate these genes although the protein mass peaks of them had been found in the MALDI-TOF trace. Furthermore, we designed primers from two known genes with different expression abundances. The PCR yields were approaching similar using the treated cDNAs as templates. These results showed that, removal of the polyA and enrichment of rare genes with DSN can meet the requirements of large-scale sequencing and discovery of new genes.

[Exploration on the relationship between pathogenesis for yin-deficiency of acute myocardial infarction and immediate prognosis as well as its mechanism].

OBJECTIVE: To study the relationship between pathogenesis for Yin-deficiency of acute myocardial infarction (AMI) and immediate prognosis as well as its neuro-endocrine mechanism. METHODS: According to the TCM standard of Syndrome Differentiation of Deficiency Syndrome, 194 patients with AMI were classified into the typical Yin-deficiency group (n = 26), the non-typical Yin-deficiency group (n = 61) and the non-Yin-deficiency group (n = 107). Their venous blood samples were collected in the morning while lying on their backs to detect plasma levels of adrenaline, noradrenaline, aldosterone, atrial natriuretic peptide (ANP), corticoid and myocardial enzymes, as well as their hospitalization days and mortality in hospitalized period were calculated and compared in the three groups, with the 30 healthy persons as control. RESULTS: Levels of serum creatine phosphokinase, creatine phosphokinase isozyme, plasma adrenaline, noradrenaline, aldosterone, hospitalization days and mortality were higher in the Yin-deficiency groups than in the non-Yin-deficiency group (P < 0.05, P < 0.01). As compared the indexes between the typical and the non-typical Yin-deficiency groups, significant difference only showed in plasma aldosterone and ANP, which was significantly higher in the former (P < 0.05, P < 0.01). Plasma corticoid level was insignificantly different between the Yin-deficieny groups. CONCLUSION: Patients with AMI of Yin-deficiency type was severer in myocardial damage, with longer hospitalization period and higher mortality, it is probably due to the hyper-activated sympathetic-adrenaline system and strengthened activity of aldosterone in them.