Developmental validation of a microRNA panel using quadratic discriminant analysis for the classification of seven forensically relevant body fluids

Rhodes, Ciara; Lewis, Carolyn; Szekely, Jennifer; Campbell, Annabelle; Creighton, Mary-Randall A.; Boone, Edward L.; Seashols‐Williams, Sarah

doi:10.1016/j.fsigen.2022.102692

Cited by 11 publications

(13 citation statements)

References 24 publications

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“…Accordingly, we attempted to investigate the performance of this combination system in mixed samples. As miRNAs distinguish body fluids by their relative expression differences, the ability of miRNAs to distinguish tissue types of the mixed body fluid samples is still in the rudimentary stage [17]. As might be expected, only mRNA markers were identified in the different mixtures of two or three kinds of body fluids, and mixtures were poorly predicted using miRNA markers (Table S7), which is consistent with previous reports [17].…”

Section: Resultssupporting

confidence: 90%

“…HBB, a globulin of hemoglobin, plays a key role in red blood cells (erythrocytes are approximately 700 times richer than white blood cells) [8]. In general, it is sometimes difficult to distinguish body fluid types, especially SA, VA, and MB, only using the relative expressions of miRNAs, which are consistent with previous reports [17]. For example, miR205-5p is an epithelium-specific marker [29,30].…”

Section: Discussionsupporting

confidence: 78%

“…As miRNAs distinguish body fluids by their relative expression differences, the ability of miRNAs to distinguish tissue types of the mixed body fluid samples is still in the rudimentary stage [17]. As might be expected, only mRNA markers were identified in the different mixtures of two or three kinds of body fluids, and mixtures were poorly predicted using miRNA markers (Table S7), which is consistent with previous reports [17]. The above results suggest there are no mRNA markers dropouts for different body fluid mixtures in the combined mRNA and miRNA system.…”

Section: 6mentioning

confidence: 99%

“…The miRNAs have been proven to be more stable than mRNA in many cases, especially in the humidity and ultraviolet radiation environments [14,16], due to their small sizes and lack of the poly-A tail. Nonetheless, previous studies have shown that miRNAs are not as specific and accurate as mRNAs in mixed-body fluid samples [17]. If mRNAs and miR-NAs in a sample can be detected by a combination system at the same time, the advantages of these two kinds of RNAs can be combined while avoiding their shortcomings.…”

Section: Introductionmentioning

confidence: 99%

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Forensic validation of a combined analysis of mRNA and miRNA markers for precise tissue origin inferences of five kinds of body fluids by RT‐qPCR

Chen,

Xu,

Zhu

2023

Electrophoresis

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Correctly inferring the tissue origin types of forensic‐relevant body fluids left at a crime scene is beneficial for reconstructing a crime scene. However, it is still a challenge to accurately identify different kinds of body fluids at a crime scene. Shorter sequence length and anti‐degradation microRNA (miRNA) can be used to infer the tissue sources of biological fluid traces, but a limited number of miRNAs are tissue specific. The application of messenger RNA (mRNA) has been confirmed by different studies based on its high tissue specificity. According to the differential expression features of mRNA or miRNA in forensically relevant body fluids, this study developed a simultaneously reversed mRNA and miRNA system and then used these two types of RNAs for the determinations of five common kinds of body fluids. Compared with previously reported single kind of mRNA or miRNA assay, the combined mRNA and miRNA system showed good advantages for human body fluid identifications, especially it could be applied in mixed samples. In conclusion, the obtained results indicated that this combined mRNA and miRNA system might provide a scientific and accurate reference for body fluid identifications.

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Section: Resultssupporting

confidence: 90%

Section: Discussionsupporting

confidence: 78%

Section: 6mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Forensic validation of a combined analysis of mRNA and miRNA markers for precise tissue origin inferences of five kinds of body fluids by RT‐qPCR

Chen,

Xu,

Zhu

2023

Electrophoresis

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“…Therefore, it is proposed in this study to expand the previously reported microbial 16S triplex to incorporate a miRNA reverse transcription‐qPCR (RT‐qPCR) duplex that can now differentiate between vaginal/menstrual secretions, feces, saliva, blood, and semen at the quantification step of the forensic DNA workflow [17]. Our previous work on body fluid classification using miRNAs had previously identified and validated miR‐891a and let‐7g as being useful as markers for distinguishing blood and semen, and as an internal control, respectively [46].…”

Section: Introductionmentioning

confidence: 99%

A combined molecular approach utilizing microbial DNA and microRNAs in a qPCR multiplex for the classification of five forensically relevant body fluids

Lewis,

Seashols‐Williams

2023

Journal of Forensic Sciences

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Body fluid identification is an essential step in the forensic biology workflow that can assist DNA analysts in determining where to collect DNA evidence. Current presumptive tests lack the specificity that molecular techniques can achieve; therefore, molecular methods, including microRNA (miRNA) and microbial signature characterization, have been extensively researched in the forensic community. Limitations of each method suggest combining molecular markers to increase the discrimination efficiency of multiple body fluids from a single assay. While microbial signatures have been successful in identifying fluids with high bacterial abundances, microRNAs have shown promise in fluids with low microbial abundance (blood and semen). This project synergized the benefits of microRNAs and microbial DNA to identify multiple body fluids using DNA extracts. A reverse transcription (RT)‐qPCR duplex targeting miR‐891a and let‐7g was validated, and miR‐891a differential expression was significantly different between blood and semen. The miRNA duplex was incorporated into a previously reported qPCR multiplex targeting 16S rRNA genes of Lactobacillus crispatus, Bacteroides uniformis, and Streptococcus salivarius to presumptively identify vaginal/menstrual secretions, feces, and saliva, respectively. The combined classification regression tree model resulted in the presumptive classification of five body fluids with 94.6% overall accuracy, now including blood and semen identification. These results provide proof of concept that microRNAs and microbial DNA can classify multiple body fluids simultaneously at the quantification step of the current forensic DNA workflow.

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Differentiation of five forensically relevant body fluids using a small set of microRNA markers

Altmeyer,

Baumer,

Hall

2024

Electrophoresis

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In forensic investigations, identifying the type of body fluid allows for the interpretation of biological evidence at the activity level. Over the past two decades, significant research efforts have focused on developing molecular methods for this purpose. MicroRNAs (miRNAs) hold great promise due to their tissue‐specific expression, abundance, lack of splice variants, and relative stability. Although initial findings are promising, achieving consistent results across studies is still challenging, underscoring the necessity for both original and replication studies. To address this, we selected 18 miRNA candidates and tested them on 6 body fluids commonly encountered in forensic cases: peripheral blood, menstrual blood, saliva, semen, vaginal secretion, and skin. Using reverse transcription quantitative PCR analysis, we confirmed eight miRNA candidates (miR‐144‐3p, miR‐451a, miR‐205‐5p, miR‐214‐3p, miR‐888‐5p, miR‐891a‐5p, miR‐193b‐3p, miR‐1260b) with high tissue specificity and four (miR‐203a‐3p, miR‐141‐3p, miR‐200b‐3p, miR‐4286) with lesser discrimination ability but still contributing to body fluid differentiation. Through principal component analysis and hierarchical clustering, the set of 12 miRNAs successfully distinguished all body fluids, including the challenging discrimination of blood from menstrual blood and saliva from vaginal secretion. In conclusion, our results provide additional data supporting the use of a small set of miRNAs for predicting common body fluids in forensic contexts. Large population data need to be gathered to develop a body fluid prediction model and assess its accuracy.

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Developmental validation of a microRNA panel using quadratic discriminant analysis for the classification of seven forensically relevant body fluids

Cited by 11 publications

References 24 publications

Forensic validation of a combined analysis of mRNA and miRNA markers for precise tissue origin inferences of five kinds of body fluids by RT‐qPCR

Forensic validation of a combined analysis of mRNA and miRNA markers for precise tissue origin inferences of five kinds of body fluids by RT‐qPCR

A combined molecular approach utilizing microbial DNA and microRNAs in a qPCR multiplex for the classification of five forensically relevant body fluids

Differentiation of five forensically relevant body fluids using a small set of microRNA markers

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