• Users Online: 856
  • Home
  • Print this page
  • Email this page
Home About us Editorial board Ahead of print Current issue Search Archives Submit article Instructions Subscribe Contacts Login 


 
 Table of Contents  
ORIGINAL ARTICLE
Year : 2019  |  Volume : 5  |  Issue : 3  |  Page : 123-129

Degradation of β-Actin mRNA and 18S rRNA in mouse spleen cells after death


1 Key Laboratory of Forensic Genetics of Ministry of Public Security, Institute of Forensic Science, Ministry of Public Security; Collaborative Innovation Center of Judicial Civilization, China; Key Laboratory of Evidence Science (China University of Political Science and Law), Ministry of Education, Beijing, China
2 Public Security Bureau of Huaian City, Jiangsu, China
3 Collaborative Innovation Center of Judicial Civilization, China; Key Laboratory of Evidence Science (China University of Political Science and Law), Ministry of Education, Beijing, China

Date of Web Publication18-Sep-2019

Correspondence Address:
Dong Zhao
Institute of Evidence Law and Forensic Science, China University of Political Science and Law, Ministry of Education, No. 25, Xitucheng Road, Haidian District, Beijing 100088
China
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jfsm.jfsm_38_19

Rights and Permissions
  Abstract 


We observed degradation of β-actin mRNA and 18S rRNA in mouse spleen cells under constant temperature conditions in the different temperature group during postmortem intervals (PMIs) of 0–72 h. Thirty-nine mice were sacrificed by cervical dislocation and kept at constant temperatures of 10°C, 15°C, 20°C, 25°C, and 30°C. From 0 to 72 h after death, total RNA in spleen cells was extracted every 6 h. The cycle threshold (Ct) values of β-actin mRNA and 18S rRNA were obtained by real-time-quantitative polymerase chain reaction. The results showed that, under the conditions of different and constant temperatures after mouse death at 72 h, the Ct values of β-actin and 18S, Ct ratios of β-actin to 18S, and relative ratios of β-actin to 18S were significantly correlated with PMI. In addition, the relative degradation rates of β-actin and 18S appeared to change from fast to slow with the increase of temperature. By interpolation and fitting analysis of the data, we obtained a ternary quintic equation of the relationship between the change in the relative ratios and PMI, which can be used to infer PMI within a certain temperature range (10°C–30°C).

Keywords: 18S rRNA, forensic pathology, interpolation function, postmortem interval, β-actin mRNA


How to cite this article:
An Z, Li F, Zhao D. Degradation of β-Actin mRNA and 18S rRNA in mouse spleen cells after death. J Forensic Sci Med 2019;5:123-9

How to cite this URL:
An Z, Li F, Zhao D. Degradation of β-Actin mRNA and 18S rRNA in mouse spleen cells after death. J Forensic Sci Med [serial online] 2019 [cited 2019 Nov 14];5:123-9. Available from: http://www.jfsmonline.com/text.asp?2019/5/3/123/267151




  Introduction Top


It is an important and difficult task in forensic practice to infer the postmortem interval (PMI).[1] At present, the gold standard for inferring the PMI in the early postmortem period is the temperature-based nomogram method together with time of death-dependent criteria of supravital reaction, postmortem turbidity of cornea, livor mortis, and rigor mortis. There is also huge literature on chemical methods proposed for estimating the time since death but obviously not yet applied in casework. Studies have shown that the number and type of internal bacteria, ambient temperature of the body, pH of the soil in which the body is located, and even the oxygen partial pressure of the body's environment affect postmortem changes.[2],[3]

To eliminate these effects, we first need to select specimens that are relatively isolated from the external environment, such as internal organs including the liver, kidney, and spleen, and then control the environmental conditions and temperature. In recent years, with the continuous development of molecular biology, the introduction of reverse transcription-polymerase chain reaction (RT-PCR) to assess time of death has become a new research focus. RNA is easily degradable, which provides new approaches to objectively infer PMI.[4],[5] β-Actin and 18S are housekeeping genes that are widely present in various eukaryotic cells. Their sequences are highly conserved and expression is abundant.[6],[7] In this study, mouse spleen cells were analyzed over 72 h after death for degradation of β-actin mRNA and 18S rRNA to provide a reference for inferring PMI.


  Materials and Methods Top


Experimental animals and materials

Thirty-nine Kunming mice (Beijing Animal Experiment Center) were randomly divided into 13 groups (1 control group and 12 experimental groups) with three mice in each group. Mice in the control group were sacrificed by cervical dislocation, and the spleen tissues were extracted immediately. Mice in the experimental groups were sacrificed by cervical dislocation and placed in incubators (temperature control range: 0°C–60°C, temperature resolution: 0.1°C; Shanghai Yiheng) at constant temperatures of 10°C, 15°C, 20°C, 25°C, and 30°C. Spleen tissues were extracted according to the time of death (6, 12, 18, 24, 30, 36, 42, 48, 54, 60, 66, and 72 h). The spleen tissue samples were quickly frozen in liquid nitrogen and stored at − 80°C. All procedures were approved by the animal facility and ethics board of our institution.

RNA extraction and purification

For RNA extraction, 0.2 g tissue was thoroughly ground in liquid nitrogen, mixed with 1 ml trizol (invitrogen), vortexed for 10 s, and incubated at 65°C for 10 min to completely lyse the cells. The sample was centrifuged at 10,000 ×g for 10 min at low temperature. The precipitate was discarded, and the supernatant was transferred to a new tube, combined with 0.2 ml chloroform, shaken vigorously for 30 s, incubated at room temperature for 3 min, and then centrifuged at 10,000 ×g for 10 min. The supernatant was transferred to a new tube, and then 0.5 ml isopropanol was added, mixed by inversion, and incubated at room temperature for 5 min. After centrifuging at 10,000 ×g for 10 min at low temperature, the supernatant was removed and 1 ml of 70% ethanol (Diethylpyrocarbonate [DEPC]) was added, followed by vigorous vortexing and then centrifugation at 7500 ×g for 5 min at low temperature. After removing the supernatant, this step was repeated two to three times. The pellet was then solubilized in 100 μl DEPC-treated H2O.

For RNA purification, 350 μl BB12 (all gold ER701) was added to the 100 μl of extracted RNA and mixed by vortexing. After addition of 900 μl absolute ethanol and mixing by inversion, the solution was transferred to a spin column and centrifuged at 10,000 ×g for 30 s twice. After adding 80 μl deoxyribonuclease I working solution (all gold ER401) and incubation at room temperature for 15 min. 500 μl WB12 (all gold ER701) was added, followed by centrifugation at 10,000 ×g for 30 s. Then, the effluent was discarded, and this step was repeated once. After centrifugation at 10,000 ×g for 2 min to completely remove residual ethanol, 20 μl RNA solution was added to the spin column, incubated for 2 min at room temperature, and then centrifuged at 10,000 ×g for 1 min to elute the RNA.

Reverse transcription and quantitative polymerase chain reaction

For reverse transcription, we used a TaqMan Gold RT-PCR kit (Applied Biosystems). The reaction mixture consisted of 0.5 μg purified RNA, 1 × TaqMan buffer, 5.5 mM magnesium chloride, 500 μM dNTPs, 1.25 U/μl MultiScribe Reverse Transcriptase, 0.4 U/μl RNase inhibitor, and 2.5 μM oligo d (T) 16. Reaction conditions were 25°C for 10 min, 48°C for 30 min, and then 95°C for 5 min.

Real-time quantitative detection of gene expression was performed using TaqMan Universal PCR Master Mix and 18S rRNA reagents (Applied Biosystems) or β-actin mRNA reagents (Applied Biosystems) and a Bio-Rad CFX96 Real-Time instrument. The reaction mixture consisted of 1 × TaqMan Universal PCR master mix, 50 nM 18S or β-actin primers, 100 nM 18S or β-actin probes, and 5 ng reverse-transcribed product. Reaction conditions were 50°C for 2 min, 95°C for 10 min, 95°C for 15 s, and 60°C for 1 min (40 cycles).

Analysis of gene expression data

For each group of three samples, the average Ct value was obtained: (1) Excel statistical software version 15.27 (161010) was used for regression analysis of data from each group; (2) We compared the Ct values of β-actin mRNA and 18S rRNA at different time points, and then regression analysis was made between the Ct ratio and PMI; (3) The fitting Ct values of the regression equation of β-actin mRNA was subtracted from the fitting Ct value of 18S rRNA in the same sample to obtain the ΔCt value; (4) The ΔCt value of each PMI group was subtracted from the ΔCt value of control group to obtain the ΔΔC value; (5) The 2-ΔΔCt value indicated the relative expression level of β-actin mRNA to 18S rRNA in the same sample, which were plotted as an exponential trend graph and analyzed by Excel software; and (6) Metlab16.0 (METLAB Inc., Natick, MA) was used for interpolation analysis and equation fitting.


  Results Top


Cycle threshold values of amplification curves of β-actin mRNA and 18S rRNA in the different temperature groups

To a certain extent, the Ct value reflects the amount of cDNA template in the PCR amplification. A small Ct value indicates that the template quantity is high and vice versa. The results showed that the Ct value of β-actin mRNA was consistently higher than that of 18S rRNA over the entire time period in all temperature groups. During prolongation of PMI, the Ct values of β-actin mRNA and 18S rRNA showed increasing trends [Table 1] and [Table 2].
Table 1: Cycle threshold values of β-actin mRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0-72 h

Click here to view
Table 2: Cycle threshold values of 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0-72 h

Click here to view


A linear trend graph of the β-actin mRNA and 18S rRNA Ct values was plotted and analyzed by Excel software. Then, regression equations for the different temperature groups were obtained as follows: for β-actin mRNA, 10°C temperature group: y = 0.0567x + 21.795, R2 = 0.5161; 15°C temperature group: y = 0.0435x + 22.39, R2 = 0.61115; 20°C temperature group: y = 0.1116x + 22.252, R2 = 0.86278; 25°C temperature group: y = 0.123x + 23.24, R2 = 0.84437; and 30°C temperature group: y = 0.1057x + 24.788, R2 = 0.56331. In 18S rRNA, 10°C temperature group: y = 0.0448x + 9.4931, R2 = 0.91099; 15°C temperature group: y = 0.0401x + 9.3468, R2 = 0.65977; 20°C temperature group: y = 0.1324x + 8.2192, R2 = 0.88254; 25°C temperature group: y = 0.1599x + 9.1605, R2 = 0.94006; and 30°C temperature group: y = 0.1614x + 10.577, R2 = 0.83124. The results showed that the Ct values of β-actin mRNA and 18S rRNA increased linearly with PMI at various time points in the different temperature groups and both had a strong positive correlation with PMI [Figure 1] and [Figure 2].
Figure 1: Changing trends of cycle threshold values of β-actin mRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0–72 h

Click here to view
Figure 2: Changing trends of cycle threshold values of 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0–72 h

Click here to view


Angle degree of the linear equation of β-actin mRNA and 18S rRNA in the different temperature groups

Based on the slope values obtained from the linear regression equations of β-actin mRNA and 18S rRNA in the different temperature groups, we calculated the angle degree of the linear equation of β-actin mRNA and 18S rRNA in the different temperature groups, which showed a gradual decline (from positive to negative) from the low temperature group to the high temperature group [Table 3]. The results indicated that the rate of relative degradation of β-actin mRNA to 18S rRNA changed from fast to slow with the increase of temperature.
Table 3: Angle degree of the linear equation of β-actin mRNA and 18S rRNA in the different temperature groups

Click here to view


The linear trend plots of the angle degree of the linear equation of β-actin mRNA and 18S rRNA in the different temperature groups were plotted. After correlation analysis by Excel software, the regression equation was obtained: Y = −0.1981x + 2.8605, R2 = 0.98581. The results showed that the angle degree decreased linearly with increasing temperature, and there was a strong negative correlation between the angle degree and temperature [Figure 3].
Figure 3: Changing trend of the angle degree of the linear equation of β-actin mRNA and 18S rRNA in the different temperature groups

Click here to view


Average cycle threshold ratios of β-actin mRNA and 18S rRNA

We compared β-actin mRNAs of the different temperature groups at various time points with the average Ct value of 18S rRNA. The results showed that the Ct ratio of β-actin mRNA to 18S rRNA over the entire time period showed a downward trend with PMI [Table 4].
Table 4: Cycle threshold ratios of β-actin mRNA to 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0-72 h

Click here to view


The linearity trend plots of Ct ratios of β-actin mRNA to 18S rRNA were plotted and analyzed by Excel software. The regression equations for different temperature groups were obtained as follows: among them, 10°C temperature group: Y = −0.0039x + 2.2975, R2 = 0.35376; 15°C temperature group: Y = −0.0042x + 2.3888, R2 = 0.28416; 20°C temperature group: Y = −0.0123x + 2.5366, R2 = 0.82605; 25°C temperature group: Y = −0.0135x + 2.4192, R2 = 0.90135; and 30°C temperature group: Y = −0.0145x + 2.3534, R2 = 0.75878. These results showed that the Ct ratio of β-actin mRNA to 18S rRNA decreased linearly with PMI at various time points in the different temperature groups [Figure 4].
Figure 4: Changing trend of cycle threshold ratios of β-actin mRNA to 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0-72 h

Click here to view


Relative expression level (relative ratio) of β-actin mRNA to 18S rRNA

To more intuitively reflect the correlation between the relative amount of the two kinds of RNA and PMI, and reduce the error caused by human factors, the fitting Ct values of the regression equation of β-actin mRNA was subtracted from the fitting Ct value of 18S rRNA in the same sample to obtain the ΔCt value. The ΔCt value of each PMI group was subtracted from the ΔCt value of control group to obtain the ΔΔC value. Moreover, the 2− ΔΔ Ct was obtained as the relative expression level of β-actin mRNA to 18S rRNA in the sample.

The results showed that the relative ratios of β-actin mRNA to 18S rRNA in 10°C and 15°C temperature groups decreased exponentially, whereas the relative ratios in 20°C, 25°C, and 30°C temperature groups showed an exponential increase with the prolongation of PMI [Table 5].
Table 5: Relative expression levels of β-actin mRNA to 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0-72 h

Click here to view


The relative ratios of β-actin mRNA to 18S rRNA were plotted as an exponential trend graph and analyzed by Excel software. Regression equations of the different temperature groups were obtained as follows: among them, 10°C temperature group: y = 1.0013e −0.008x, R2 = 0.99963; 15°C temperature group: y = 1.002e −0.002x, R2 = 0.99608; 20°C temperature group: y = 1.0001e 0.0143x, R2 = 0.99998; 25°C temperature group: y = 1.0009e 0.0256x, R2 = 0.99997; and 30°C temperature group: y = 1.0005e 0.0386x, R2 = 0.99998. The results showed that, with prolongation of PMI, the relative ratio of β-actin mRNA to 18S rRNA in 10°C and 15°C temperature groups showed an exponentially decreasing trend (from an obvious to slow decrease), whereas the relative ratios in 20°C, 25°C, and 30°C temperature groups showed an exponentially upward trend (from slow, faster, and to more pronounced). The changing trend of the relative ratio was significantly related to temperature, indicating that the relative rate of degradation of β-actin mRNA to 18S rRNA showed changes from fast to slow with increasing temperature [Figure 5].
Figure 5: Changing trend of the relative expression of β-actin mRNA to 18S rRNA in mouse spleen cells in the different temperature groups during postmortem intervals of 0–72 h

Click here to view


According to the interpolation and fitting analysis of the obtained data, a ternary quintic equation of the relationship between the change in relative ratios and PMI was obtained within a certain temperature range (10°C–30°C): F(x, y) = p00 + p10*x + p01*y + p20*x^2 + p11*x*y + p02*y^2 + p30*x^3 + p21*x^2*y + p12*x*y^2 + p03*y^3 + p40*x^4 + p31*x^3*y + p22*x^2*y^2 + p13*x*y^3 + p04*y^4 + p50*x^5 + p41*x^4*y + p32*x^3*y^2 + p23*x^2*y^3 + p14*x*y^4 + p05*y^5; Coefficients (with 95% confidence interval): P00 = −50.64 (−84.22, −17.05), p10 = 68.99 (10.58, 127.4), p01 = 3.595 (0.6697, 6.52), p20 = 0.7321 (−36.54, 38), p11 = −4.175 (−8.275, −0.07547), p02 = −0.08337 (−0.2217, 0.05498), p30 = 2.446 (−8.184, 13.08), p21 = −0.8062 (−3.339, 1.727), p12 = 0.1575 (0.004296, 0.3107), p03 = 3.005e-05 (−0.003726, 0.003787), p40 = −1.414 (−4.385, 1.557), p31 = 0.2276 (−0.6351, 1.09), p22 = 0.003028 (−0.0664, 0.07245), p13 = −0.002222 (−0.00502, 0.0005758), p04 = 1.757e-05 (−3.697e-05, 7.211e-05), p50 = −0.03077 (−0.09479, 0.03325), p41 = 0.03002 (−0.03536, 0.0954), p32 = −0.004461 (−0.01778, 0.008857), p23 = 0.0001188 (−0.0007456, 0.0009831), p14 = 9.812e-06 (−1.281e-05, 3.244e-05), p05 = −1.345e-07 (−4.394e-07, 1.704e-07); R-squared: 0.9034.

A curved surface diagram of the relationship between temperature, relative ratio, and PMI is shown in [Figure 6].
Figure 6: Curved surface diagram of the relationship between the change in relative ratios and postmortem interval within a certain temperature range (10°C–30°C)

Click here to view



  Discussion Top


β-actin mRNA and 18S rRNA degradation and postmortem interval

RNA includes mRNA, tRNA, and rRNA. Different types of RNA may degrade at different rates, which leads to changes in the ratio of RNA types over time.[8] To test the hypothesis that different types of RNAs decay at different rates in vivo, we compared the relative stability of an mRNA to an rRNA.

Mature β-actin mRNA is mainly contained in the cytoplasm, which is easily degraded and has a short half-life. Electron microscopic observation shows that the polymer of β-actin mRNA is not very compact, there is a clear gap in the middle, so it is easily interfered with by the external environment and its stability is poor.[9] 18S rRNA is ubiquitous in ribosomal protein complexes and has thousands of copies in each cell. Ribosomal proteins can separate RNase and other chemical factors, leaving 18S rRNA relatively stable when cells are not damaged and ribosomes are not dissolved.[10] Therefore, both β-actin mRNA and 18S rRNA were suitable for this study.

In this study, Excel software was used to analyze the Ct changes of β-actin mRNA and 18S rRNA in mouse spleen cells within 72 h after death in different temperature groups. Regression equations were obtained and the results showed that the Ct values of β-actin mRNA were always higher than those of 18S rRNA in all temperature groups, and the Ct values of β-actin mRNA and 18S rRNA increased with PMI. The correlations between the Ct values and PMI were strong, and R2 ranged from 0.5161 to 0.94006, all of which were > 0.25. Among them, β-actin mRNA had the best correlation with PMI in 20°C and 25°C temperature groups (R2 = 0.86278 and R2 = 0.84437), and 18S rRNA had the best correlation with PMI in 10°C and 25°C temperature groups (R2 = 0.91099 and R2 = 0.94006). These results confirmed that the Ct values of β-actin mRNA and 18S rRNA increased linearly with prolongation of PMI at various time points in the different temperature groups, and both of them had a strong positive correlation with PMI.

Angle degree of the linear equation of β-actin mRNA and 18S rRNA with temperatures

Based on the regression equations of the Ct values of β-actin mRNA and 18S rRNA in the above different temperature groups, we can see that the linear slopes of different temperature groups are different, which also confirms the difference in the degradation rate of β-actin mRNA and 18S rRNA after death in mice.

With the increase of temperature from 10°C to 30°C, the angle degree showed a linear downward trend, and there was a strong negative correlation between the angle degree and temperature. It also showed that the degradation rate of β-actin mRNA relative to 18S rRNA changed from fast to slow with the increase in temperature, indicating that mRNA did not always degrade more rapidly than rRNA.

Average Ct ratio of β-actin mRNA to 18S rRNA with postmortem interval

Both RNAs were attenuated in a consistent and predictable manner. By detecting the RNA ratio, the analysis was independent of the sample amount, and detection required only a relatively small sample to be successfully processed. Species specificity of primers and probes helps to eliminate false signals due to contamination. All experimental manipulations occurred simultaneously for both target RNAs in the same tube, which eliminated potential problems such as pipetting errors and differences in enzyme efficiency. In this study, the results showed that the average Ct ratio of β-actin mRNA to 18S rRNA at the various time points in the different temperature groups had a linear downward trend with PMI, and the correlation between them was strong. This result is consistent with the results of Anderson reported that the Ct ratio of 18S rRNA and β-actin mRNA gradually increased with the prolongation of blood mark formation within 150 days at room temperature.[8]

In addition, R2 ranged from 0.28416 to 0.90135, all of which were >0.25. Among them, the average Ct ratios had the best correlation with PMI in 20°C and 25°C temperature groups (R2 = 0.82605 and R2 = 0.90135).

Relative expression level (relative ratio) of β-actin mRNA to 18S rRNA

Two of the most common methods to analyze real-time quantitative PCR data are absolute and relative quantifications. The purpose of absolute quantification is to determine the number of molecules of the gene of interest in the sample, the so-called “copy number.” The purpose of relative quantification is to determine the relative proportions of genes of interest in two or more samples without knowing their copy number in each sample. In the process of obtaining materials, differing factors such as methods and measurement tools often lead to deviations in the absolute quantification of samples. Therefore, the finding reliable parameter indexes for relative quantification have become the preferred method to infer PMI. The 2− ΔΔ Ct method is a convenient method to analyze relative changes in gene expression through real-time-quantitative PCR.

In this study, all temperature groups had a relatively high correlation between PMI and the relative ratio. R2 ranged from 0.99608 to 0.99998, all of which were much larger than 0.25. Among them, with the prolongation of PMI, the relative ratio of β-actin mRNA to 18S rRNA in 10°C and 15°C temperature groups showed an exponentially decreasing trend (from an obvious to slow decrease), whereas the relative ratios in 20°C, 25°C, and 30°C temperature groups showed an exponential upward trend (from slow, faster, and to more pronounced). The changing trend of the relative ratio was significantly related to temperature, indicating that the relative rate showed changes from fast to slow with increasing temperature. Based on these results, PMI can be well inferred by the temperature change. The method also successfully avoided the deficits in the absolute quantitative method, and the results were more reasonable.

Limitations of this study

We used relative quantitative methods to detect the degradation of β-actin mRNA and 18S rRNA, but we did not detect the integrity of RNA, which can be reflected by the RNA integrity number (RIN). RIN has become the standard method to measure RNA integrity.[11] In addition, the temperature was controlled precisely, which is different from the temperature change of real environmental conditions. Temperature is the main factor influencing PMI inference. In the next step to infer PMI, RIN indicators should be used to assign integrity values to RNA measurements, and it is necessary to operate under natural environmental temperature. Furthermore, this study only explored the degradation of β-actin mRNA and 18S rRNA in spleen cells of the different temperature groups at constant temperature without considering other external environmental factors.


  Conclusion Top


Under the conditions of different and constant temperatures after mouse death, the Ct values of β-actin and 18S, Ct ratios of β-actin to 18S, and relative ratios of β-actin to 18S were significantly correlated with PMI. We also obtained a ternary quintic equation of the relationship between the change in relative ratios and PMI, which can be used to infer PMI within a certain temperature range (10°C–30°C).

Financial support and sponsorship

This work was supported by the Open Project of Key Laboratory of Forensic Genetics, Ministry of Public Security (No. 2017FGKFKT05), Beijing Natural Science Foundation (No. 7192121), and Chinese Academy of Engineering (No. 2019-XZ-31).

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Alibegović A. Cartilage: A new parameter for the determination of the postmortem interval? J Forensic Leg Med 2014;27:39-45.  Back to cited text no. 1
    
2.
Vass AA, Barshick SA, Sega G, Caton J, Skeen JT, Love JC, et al. Decomposition chemistry of human remains: A new methodology for determining the postmortem interval. J Forensic Sci 2002;47:542-53.  Back to cited text no. 2
    
3.
Forbes SL, Perrault KA. Decomposition odour profiling in the air and soil surrounding vertebrate carrion. PLoS One 2014;9:e95107.  Back to cited text no. 3
    
4.
C Zapico S, Menéndez ST, Núñez P. Cell death proteins as markers of early postmortem interval. Cell Mol Life Sci 2014;71:2957-62.  Back to cited text no. 4
    
5.
Lv YH, Ma KJ, Zhang H, He M, Zhang P, Shen YW, et al. A time course study demonstrating mRNA, microRNA, 18S rRNA, and U6 snRNA changes to estimate PMI in deceased rat's spleen. J Forensic Sci 2014;59:1286-94.  Back to cited text no. 5
    
6.
Latham VM Jr., Kislauskis EH, Singer RH, Ross AF. Beta-actin mRNA localization is regulated by signal transduction mechanisms. J Cell Biol 1994;126:1211-9.  Back to cited text no. 6
    
7.
Sloma MS, Nygård O. Chemical accessibility of 18S rRNA in native ribosomal complexes: Interaction sites of mRNA, tRNA and translation factors. Biol Chem 2001;382:661-8.  Back to cited text no. 7
    
8.
Anderson S, Howard B, Hobbs GR, Bishop CP. A method for determining the age of a bloodstain. Forensic Sci Int 2005;148:37-45.  Back to cited text no. 8
    
9.
Lü YH, Li ZH, Tuo Y, Liu L, Li K, Bian J, et al. Correlation between RNA degradation patterns of rat's brain and early PMI at different temperatures. Fa Yi Xue Za Zhi 2016;32:165-70.  Back to cited text no. 9
    
10.
Sidova M, Tomankova S, Abaffy P, Kubista M, Sindelka R. Effects of post-mortem and physical degradation on RNA integrity and quality. Biomol Detect Quantif 2015;5:3-9.  Back to cited text no. 10
    
11.
Schroeder A, Mueller O, Stocker S, Salowsky R, Leiber M, Gassmann M, et al. The RIN: An RNA integrity number for assigning integrity values to RNA measurements. BMC Mol Biol 2006;7:3.  Back to cited text no. 11
    


    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]



 

Top
 
 
  Search
 
Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
Access Statistics
Email Alert *
Add to My List *
* Registration required (free)

 
  In this article
Abstract
Introduction
Materials and Me...
Results
Discussion
Conclusion
References
Article Figures
Article Tables

 Article Access Statistics
    Viewed285    
    Printed29    
    Emailed0    
    PDF Downloaded69    
    Comments [Add]    

Recommend this journal


[TAG2]
[TAG3]
[TAG4]