ABSTRACT
Objective To establish a convenient and rapid method for extracting DNA from bone.Methods Fifteen long bone samples were washed and sterilized.The skeletal fragments were obtained by electric drill,and lysed by PrepFiler Express BTATM lysis buffer.DNA was then manually extracted by silicon microbeads for further analysis.Results STR genotyping was successfully obtained in 14 out of the 15 samples,and the detection rate was 93.33％.Conclusion The method for DNA extraction from bone established in present study is convenient,quick,effective,and with a strong applicability,which is worth spreading and applying.
ABSTRACT
Myocytes in the pulmonary veins (PV) play a pivotal role in the development of paroxysmal atrial fibrillation (AF). It is therefore important to understand physiological characteristics of these cells. Studies on these cells are, however, markedly impeded by the fact that single PV myocytes are very difficult to obtain due to lack of effective isolation methods. In this study, we described a novel PV myocyte isolation method. The key aspect of this method is to establish a combination of retrograde heart perfusion (via the aorta) and anterograde PV perfusion (via the pulmonary artery). With this simultaneous perfusion method, a better perfusion of the PV myocytes can be obtained. As results, the output and viability of single myocytes isolated by simultaneous heart and PV perfusion method were increased compared with those in conventional retrograde heart perfusion method.
Subject(s)
Animals , Rabbits , Atrial Fibrillation , Cell Separation , Heart , Muscle Cells , Perfusion , Pulmonary VeinsABSTRACT
OBJECTIVE@#To analyze and discuss four methods of calculating likelihood ratio of DNA mixture.@*METHODS@#In the case with CNAS-T0757 proficiency testing in 2013, the likelihood ratios were calculated and compared among four methods, including unrestricted combinatorial method, Clayton's method, p2 principle method, and recommendations from ISFG.@*RESULTS@#The likelihood ratios were maximum by Clayton's method and recommendations from ISFO, followed by result of the unrestricted combinational method. The minimum likelihood ratio was obtained by p2 principle.@*CONCLUSION@#The unrestricted combinational method could give fUrthest consideration to both information preservation and appraiser protection.
Subject(s)
Humans , DNA/genetics , DNA Fingerprinting , Likelihood FunctionsABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression of CSN complex (COP9 signal some subunits) in the patients with acute promyelocytic leukemia (APL) and its significance in the ATRA-induced APL differentiation.</p><p><b>METHODS</b>Using the NB4 cells as a model, morphologic observation and myeloid differentiation marker CD11b detection were used to monitor ATRA-induced APL differentiation, the expression of CSN complex in cell differentiation was detected by Western blot and reverse transcription real time fluorescent quantitative PCR (RT-qPCR) method. RT-qPCR was also used to detect the relative expression level of COP9 signalosome subunits in the APL patients and remission after treatment.</p><p><b>RESULTS</b>ATRA could obviously enhance CD11b expression; the cell morphology showed obvious differentiation characteristics. During the differentiation, the expression of COP9 signalosome subunits was down-regulated by ATRA. Meanwhile, the CSN expression level in newly diagnosed APL patients was much higher than that in controls (non-leukemia) (P < 0.05). The level of CSN expression was obviously down-regulated when APL patients achieved complete remission.</p><p><b>CONCLUSION</b>The high CSN expression level in APL patients can be down-regulated by ATRA. CSN complex may have a significant effect on the pathogenesis and therapy of APL.</p>
Subject(s)
Humans , COP9 Signalosome Complex , Cell Differentiation , Cell Line, Tumor , Down-Regulation , Leukemia, Promyelocytic, Acute , Metabolism , Multiprotein Complexes , Metabolism , Peptide Hydrolases , Metabolism , Tretinoin , PharmacologyABSTRACT
OBJECTIVE@#To explore a new method in order to extract DNA from bones and teeth automatically.@*METHODS@#Samples of 33 bones and 15 teeth were acquired by freeze-mill method and manual method, respectively. DNA materials were extracted and quantified from the triturated samples by AutoMate Express forensic DNA extraction system.@*RESULTS@#DNA extraction from bones and teeth were completed in 3 hours using the AutoMate Express forensic DNA extraction system. There was no statistical difference between the two methods in the DNA concentration of bones. Both bones and teeth got the good STR typing by freeze-mill method, and the DNA concentration of teeth was higher than those by manual method.@*CONCLUSION@#AutoMate Express forensic DNA extraction system is a new method to extract DNA from bones and teeth, which can be applied in forensic practice.
Subject(s)
Humans , Automation , Bone and Bones/chemistry , DNA/isolation & purification , DNA Fingerprinting/methods , Forensic Medicine/methods , Microsatellite Repeats , Polymerase Chain Reaction , Specimen Handling/methods , Tooth/chemistryABSTRACT
With the development of molecular biology, the evidences of genetics has been used widely in forensic sciences. DNA technology has played an important role in individual identification and paternity testing, RNA technology is showing more and more wide application in prospect. This article reviews the application and progress of RNA in forensic science including estimation of postmortem interval, bloodstain age, wound age, as well as determination of cause of death and the source of body fluids.
Subject(s)
Animals , Humans , Actins/metabolism , Blood Stains , Body Fluids/metabolism , Cause of Death , Forensic Medicine/methods , Gene Expression , Genetic Markers/genetics , MicroRNAs/genetics , Polymerase Chain Reaction/methods , Postmortem Changes , RNA/genetics , RNA Stability , RNA, Messenger/metabolism , RNA, Ribosomal, 18S/geneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate the incidence of JAK2V617F gene point mutation in patients with myeloproliferatives diseases (MPD) and its clinical significance.</p><p><b>METHODS</b>Genomic DNA from bone marrow and peripheral blood cells were extracted from 68 patients with MPD. Allele specific polymerase chain reaction was used to amplify the exon 12 of JAK2 gene which harbours V617F mutation. The PCR products were identified by DNA sequencing. JAK2V617F gene point mutation and its impact on peripheral blood cells were analyzed.</p><p><b>RESULTS</b>The incidence of JAK2V617F mutation in 68 patients with MPD was 65.28 %. The positive rate of JAK2V617F point mutation was 77.77 % in patients with PV (36/59), 56.52 % in patients with ET (23/59) and 44.44 % in patients with IMF (4/9). In all groups, the incidence of JAK2V617F point mutation in bone marrow and peripheral blood were equal. Patients with JAK2V617F mutation in PV group had higher counts of white blood cell and hemoglobin in peripheral blood than patients without JAK2V617F point mutation (P <0.05). Patients with JAK2V617F mutation in ET group had higher counts of white blood cell than those without JAK2V617F mutation (P <0.05); there was no significant difference in platelet count.</p><p><b>CONCLUSION</b>JAK2V617F point mutation can affect the hematologic features, which may be of diagnostic value for MDP with negative BCR-ABL gene.</p>
Subject(s)
Adolescent , Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , Amino Acid Substitution , Base Sequence , Janus Kinase 2 , Genetics , Molecular Sequence Data , Myeloproliferative Disorders , Genetics , Point MutationABSTRACT
We isolated mouse embryonic cardiomyocytes derived from timed-pregnant females at different periods and used patch-clamp technique to investigate the muscarinic cholinergic modulation of pacemaker current I(f) in different developmental stages. In early development stage (EDS), muscarinic agonist carbachol (CCh) significantly decreased the magnitude of the pacemaker current I(f) but had no effect in late development stage (LDS). Forskolin (a direct adenylate cyclase activator) and IBMX (a non-selective phosphodiesterase inhibitor) increased I(f) in both EDS and LDS cells. Interestingly, although both forskolin and IBMX increased basal I(f), their effects on CCh-inhibited I(f) were different. Forskolin did not reverse the inhibitory action of CCh until intermediate development stage (IDS). In contrast, IBMX reversed the inhibitory action of CCh on I(f) in EDS but not in IDS. It is suggested that a decrease in intracellular cAMP is a possible mechanism for CCh to modulate I(f). During the EDS and IDS CCh controls the cytoplasmic cAMP level by different pathways: In EDS, CCh modulates I(f) possibly by activating PDE which accelerates the breakdown of cAMP, but in IDS possibly by inhibiting adenylate cyclase (AC) which then reduces the synthesis of cAMP.