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Exploring Phylogeny in the Introductory Microbiology Lab

Exploring Phylogeny in the Introductory Microbiology Lab. Mohammed Abbas (Schoolcraft College) Kuodi Jian (Metropolitan State University) Shaily Menon (Grand Valley State University) Randy Strobel (Metropolitan State University). Introduction.

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Exploring Phylogeny in the Introductory Microbiology Lab

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  1. Exploring Phylogeny in the Introductory Microbiology Lab Mohammed Abbas (Schoolcraft College) Kuodi Jian (Metropolitan State University) Shaily Menon (Grand Valley State University) Randy Strobel (Metropolitan State University)

  2. Introduction • A universal introductory lab in Microbiology involves identification of unknown microbes by students • Here, we enhance this activity by incorporating bioinformatics and phylogenetic analysis

  3. Advantages of this approach • Adds an evolutionary component to an otherwise descriptive and physiological exercise • Students can compare a qualitative taxonomy technique with a more phylogenetic technique

  4. Methods • First, students would be asked to fill in a matrix of organisms versus various morphological and physiological characteristics based on laboratory exercises done throughout the semester • Then they would use McClade to construct cladograms

  5. Methods (contd) • Then, students would harvest sequences for an enzyme or marker where sequence information is available for all the species • Next, students would construct a molecular phylogeny using the sequence data • Finally, students would compare the two phylogenies and discuss evolutionary relationships

  6. Case study • For purposes of demonstration, we chose a limited set of 7 microbes including the 3 domains (Archaea, Eukaryotes, Bacteria) • Within the bacteria we included a gram positive, gram negative, and an acid-fast organism • We chose glucokinase (hexokinase) presuming it would be a fairly ubiquitous enzyme and likely to be sequenced in these microbes [Note: Students would pick their own marker]

  7. Matrix

  8. Cladogram from McClade

  9. Amino Acid Sequences of Glucokinase for 7 Microbes >NP_645110.1 Staphylococcus aureus MYYIAIDIGGTQIKSAVIDKQLNMFDYQQISTPDNKSELITDKVYEIVTGYMKQYQLIQPVIGISSAGVV DEQKGEIVYAGPTIPNYKGTNFKRLLKSLSPYVKVKNDVNAALLGELKLHQYQAERIFCMTLGTGIGGAY KNNQGHIDNGELHKANEVGYLLYRPTENTTFEQRAATSALKKRMIAGGFTRSTHVPVLFEAAEEGDDIAK QILNEWAEDVAEGIAQIQVMYDPGLILIGGGISEQGDNLIKYIEPKVAHYLPKDYVYAPIQTTKSKNDAA LYGCLQ >NP_764788.1 Staphylococcus epidermidisMTDIILAADIGGTTCKLGIFDKDLEQLHKWSIDTDTSDHTGELLLKNIYNSFTEKIAEYKYDFNNVVGVGIGVPGPVDFDTGVVYGAVNLHWPDSVNVREIFKQYVNCPVYVDNDANVAALGEKHKGAGEGADDVVAITLGTGLGGGIISNGEIVHGHNGSGAEIGHLRADFDQRFQCNCGKSGCIETVASATGVVNLVNFYYPKLTFKSSILQLIKDNQVTAKAVFDAAKAGDQFCIFITEKVANYIGYLCSIISVTSNPKYIVLGGGMSTAGLILIENIKTEYRNLTFTPAQNNTEIVQAKLGNDAGITGAAGLIKTYIIDKEGAK >NP_416889.1 Escherichia coli K12MTKYALVGDVGGTNARLALCDIASGEISQAKTYSGLDYPSLEAVIRVYLEEHKVEVKDGCIAIACPITGDWVAMTNHTWAFSIAEMKKNLGFSHLEIINDFTAVSMAIPMLKKEHLIQFGGAEPVEGKPIAVYGAGTGLGVAHLVHVDKRWVSLPGEGGHVDFAPNSEEEAIILEILRAEIGHVSAERVLSGPGLVNLYRAIVKADNRLPENLKPKDITERALADSCTDCRRALSLFCVIMGRFGGNLALNLGTFGGVFIAGGIVPRFLEFFKASGFRAAFEDKGRFKEYVHDIPVYLIVHDNPGLLGSGAHLRQTLGHIL >AAL17915.1 Mycobacterium smegmatisIVRRAEKEFPVTGDAAVLRETVQDTVFGCLQGSPLCGVGVASAGLVDHTTGVLRRIDDQPGFSGYPIAASLTRLLGCPVFVDHRARLQVLGDRWFGHGIGRASFASVATGDTLGLGILFEGKVIAPLGGRSGAHITVALGGRRCSCGNLGCWKTIATSTWLRQEAATRGLGPIDGVGDLIAAARGDTGARRLMTDYADNLAVGLSSVQHLLAPGLYIVHGDAAAGGGTFLGRIEERLRAVSEWADFAERPRVVAADTAADDVALLGGAGLVLSRLSGQ >YP_005657.1 Thermus thermophilusMKVVGLDLGGTKIAAGVFDGKRLLSKVVVPTPKEGGERVAEALAEAAERAEREAGVRGEAIGLGTPGPLDFRRGVIRFAPNIPGVQDFPIRRILEEATGRPVFLENDANAAALAEHHLGAAQGEESSLYLTVSTGIGGGVVLGGRVLRGERGQGGELGHLTLLPGGPACGCGLEGCLEALAAGRALERDATYAFQRPVDTRELFRLFQAGDPKAERLVLQAARYVGIGLASLVKAFDPGVVVLGGGVALNAPEGYWEALLEAYRRYLQGWEAPPLRRARLGAEAGLLGAALTAYLEVKDGSG >NP_009890.1 Saccharomyces cerevisiaeMSFDDLHKATERAVIQAVDQICDDFEVTPEKLDELTAYFIEQMEKGLAPPKEGHTLASDKGLPMIPAFVTGSPNGTERGVLLAADLGGTNFRICSVNLHGDHTFSMEQMKSKIPDDLLDDENVTSDDLFGFLARRTLAFMKKYHPDELAKGKDAKPMKLGFTFSYPVDQTSLNSGTLIRWTKGFRIADTVGKDVVQLYQEQLSAQGMPMIKVVALTNDTVGTYLSHCYTSDNTDSMTSGEISEPVIGCIFGTGTNGCYMEEINKITKLPQELRDKLIKEGKTHMIINVEWGSFDNELKHLPTTKYDVVIDQKLSTNPGFHLFEKRVSGMFLGEVLRNILVDLHSQGLLLQQYRSKEQLPRHLTTPFQLSSEVLSHIEIDDSTGLRETELSLLQSLRLPTTPTERVQIQKLVRAISRRSAYLAAVPLAAILIKTNALNKRYHGEVEIGCDGSVVEYYPGFRSMLRHALALSPLGAEGERKVHLKIAKDGSGVGAALCALVA >NP_251883.1 [Pseudomonas aeruginosa PAO1]MNNDNKRSAGGLGLVGDIGGTNARFALWRGQRLESIEVLACADYPRPELAVRDYLARIGESVANIDSVCLACAGPVGAADFRFTNNHWVINRAAFREELGLDHLLLVNDFSTMAWAASRLGADELVQVRAGSAQADRARLIIGPGTGLGVGSLLPLGGGRWEVLPCEGGHVDLPVTSPRDFALWQGLQARYGHVSAERALSGNGLLALYEISCALDGVAVRASSAAEVGALAMAGDAQADAVLEHFFLWLARVAGNAVLTVGALGGVYITGGIVPRFLERFIASGFAEAFASRGKTSGAYLQDVPVWVMTAEHPGLLGAGVALQQALDAEG 45

  10. Molecular Phylogeny Default output from a CLUSTALW alignment of the 7 microbe sequences

  11. Conclusions • Plans are to implement this exercise this fall and update the Bioquest web page with results, modifications, and suggestions for use by others • Other groups have indicated plans to steal and implement our idea 

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