1. Today HK Tree assignment *.ppt Reports, snail/parasite or fungus
Standing
Genbank submissions
Course feedback
*.ppt
Reports, snail/parasite or fungus

3. 100 65
100
COI
100 78 81 81

4. Assignment Questions
1 Is your phylogenetic analysis consistent with current snail taxonomy?
Yes, the tree reflects the three families of Hygrophyla
(P= Physidae, AP= Ancyloplanorbidae, L= Lymnaeidae), relative to outgroup,
with grouping of species previously assigned to these families
2 Do the results support the species-level identification of our experimental samples obtained in class based on 16S sequence data?
Yes,
COIP1E19, P1E; COIP2E19, P2E; CO319SIll, P3S group among Physidae
COIL1S19, L1S groups with Pseudosuccinea columella
COA19Ill, A groups with Laevapex fuscus

5. STANDING

6. SNAIL AND PARASITES BIOLOGY
DNA
“identity, possibilities”
phylogenetics
CTAB/DNAzol
gel electrophoresis
nanodrop spec
Illumina (full)
genome sequencing
PCR rDNA/mito
Qubit Fluorometry
Covaris fragmentation
Ampure (fragment collection)
Kapa DNA library preparation kit
Pippin size selection
QC Bioanalyzer, Qubit, qPCR
Illumina run
TA cloning, B/W screening
electrophoresis
Pure Link plasmid extraction Restriction digests
direct sequencing
M13 sequencing
Sequence ID (BLAST)
editing
Primer design, walking
Galaxy
QC Data file
(MT) genome assembly
Mitos, manual annotation
Gene annotation
Phylogenetics
GenBank submission

7. From: gb-admin@ncbi. nlm. nih. gov <gb-admin@ncbi. nlm. nih
From: Sent: Tuesday, November 19, :12 AM To: Coenraad Adema Subject: GenBank Submissions grp  
Dear Dr. Adema: Regarding: ========================================================= A errant mouseclick led to a premature submission. The annotation is incomplete at this time. I would like to continue annotation for this sequence before it is evaluated. So I do not want to invalidate the submission ID but I do need to add more information to the file ========================================================= Please clarify what additional information is to be added. If there are just a few, small edits - we can make the changes for you. However, if there are large edits, we will need you to resubmit the file. For Bankit submissions, start a new submission and use the same address as your original submission. You will receive a new Bankit number. Do not use the Bankit Update page to resubmit. Please also include one of these notes to help track your submission:         resubmission of bankitxxxxxx         resubmission of grp ###### (from this )         Send your response to:  and your resubmitted sequence(s) through Bankit If we do not hear from you by Dec  3, 2019, all of your submission(s) will be deleted from the processing queue. Thank you for your attention. We look forward to hearing from you so we may continue processing your submission in a timely manner. For your reference, please find your preliminary flatfiles below with the information we currently have. Please reply using the current Subject line. Sincerely, Lori Black, PhD GenBank Submissions Staff Bethesda, Maryland USA ******************************************************************* (for replies/updates to records in GenBank) (for general questions regarding GenBank) *******************************************************************
preliminary GenBank flatfiles: LOCUS       COI                      655 bp    DNA     linear   INV 18-NOV-2019 DEFINITION  Pseudosuccinea columella mitochondrion. ACCESSION   VERSION KEYWORDS    . SOURCE      mitochondrion Pseudosuccinea columella   ORGANISM  Pseudosuccinea columella             Eukaryota; Metazoa; Lophotrochozoa; Mollusca; Gastropoda;             Heterobranchia; Euthyneura; Panpulmonata; Hygrophila; Lymnaeoidea;             Lymnaeidae; Pseudosuccinea. REFERENCE   1  (bases 1 to 655)   AUTHORS   Adema,C.M.   TITLE     Hygrophilid snails and Trematode parasites from NM   JOURNAL   Unpublished REFERENCE   2  (bases 1 to 655)   AUTHORS   Hamm,P.S. and Badgett,J.E.   TITLE     Direct Submission   JOURNAL   Submitted (18-NOV-2019) Biology, U of New Mexico, 1 U of New             Mexico, albuquerque, NM 87131, US COMMENT     Bankit Comment: LocalID:COI.             Bankit Comment: BankIt                         ##Assembly-Data-START##             Sequencing Technology :: Sanger dideoxy sequencing             ##Assembly-Data-END## FEATURES             Location/Qualifiers      source                               /organism="Pseudosuccinea columella"                      /organelle="mitochondrion"                      /mol_type="genomic DNA"                      /db_xref="taxon:31228"                      /country="USA: Albuquerque NM"                      /lat_lon=" N W"                      /collection_date="23-Aug-2019"                      /PCR_primers="fwd_name: lco1490, fwd_seq:                      ggtcaacaaatcataaagatattgg, rev_name: hc02198, rev_seq:                      taaacttcagggtgaccaaaaaatca"      gene            <1..>655                      /gene="COI"      CDS             <1..>655                      /gene="COI"                      /codon_start=2                      /transl_table=5                      /product="cytochrome oxidase subunit I"                      /translation="TLYMVFGVWCGLVGTGLSLLIRLELGTSMVLIDEHFYNVIVTAH                      AFVMIFFMVMPMMIGGFGNWMVPLLIGAPDMSFPRMNNMSFWLLPPSFILLLCSSMVE                      GGVGTGWTVYPPLSGPIAHGGSSVDLAIFSLHLAGLSSILGAINFITTIFNMRSPGIT                      LERMSLFVWSVLVTAFLLLLSLPVLAGAITMLLTDRNFNTTFFDPAGGGDPILYQHLF                      " BASE COUNT      173 a     93 c    112 g    277 t ORIGIN              1 tactttatat atagtttttg gtgtatgatg tggtttagtt ggaacaggtt tgtccttatt        61 aattcgttta gaattaggta catctatagt tttaattgac gagcattttt ataatgtaat       121 tgttactgct catgcttttg ttataatttt tttcatagtt ataccaataa taattggagg       181 gtttggaaat tgaatagttc cacttctcat tggtgctcca gatataagat ttcctcgtat       241 aaataatata agattttgat tactaccacc ttcgtttatt ctcttacttt gctctagaat       301 agtagaaggt ggggtaggta ctggatgaac agtttaccca ccattgagtg gacctattgc       361 tcatggtgga tcttctgttg atttagctat tttttcttta catttagccg gtttatccag       421 gattttagga gcaattaatt ttattactac aatttttaat atacgatctc caggtattac       481 attagaacga ataagattat ttgtatgatc tgtattagtt acagcttttt tgcttctttt       541 atctttacca gtacttgcag gggcaattac aatgctttta acagatcgaa attttaatac       601 cacttttttt gatcctgctg gaggtggtga tcctatttta tatcaacatc ttttt //

9. *.ppt

10. Content Final Report Snails and Parasites
Intro Trematodes cause infectious diseases of human and animals. Characterization of snails and their trematodes present at Shady Lakes. (Potential risks for…) Methods Snail collection, inspection for parasites, broad ID on morphology. DNA extraction, Ancylid, new observation no parasite observed, but investigated to evaluate ID, potential for parasite transmission, conserved primer PCR amplification, sequencing Sanger (direct/cloning, primer walking) Contig assembly COI, 16S, 28S. Illumina sequencing, MITOBIM assembly, MITOS annotation. BLAST and Phylogenetic assembly for ID. Sequence submission to GenBank. Results Snails collected, types of parasites observed initial ID. Sequences obtained from snails and parasites. Illumina data, assembly of complete mt genomes snails and parasites. Data submitted to GenBank. BLAST and Phylogenetics of 16S and COI ID snails as v,w,x,y,z (show a tree); 28S or Mt genomes ID parasites a,b,c,d of the snails, respectively. Discussion Presence of snails and vertebrates great potential for trematode life cycles. L, P, AP can transmit several diseases. Parasite-snail associations reported previously? ID Sanger versus Illumina more specific? Specific or more general associations? Types of disease in vertebrate hosts. Risks at Shady lakes. …. Literature cited, figure/table etc.

11. Content Final Report - Lung Fungi
Intro Until very recently, healthy mammalian lungs were thought to represent a sterile environment. It is now known that they possess a true microbiome. Here, we are addressing the question of the fungal component of the lung microbiome (mycobiome) Methods a. Animals were collected in California by James Patton, Univ of California Berkeley (see spreadsheet). b. Lung fragments were plated onto YG agar medium with streptomycin (10 mg/liter) and chloramphenicol (50 mg/liter) to prevent bacterial contamination. YG agar (per liter) has 10 g yeast extract, 20 g glucose and 15 g bacto agar. c. DNA was extracted from fungal colonies using the CTAB procedure d. Fungal ITS sequences were amplified using primers ITS1-F and ITS4 e. ITS PCR fragments were sequenced (2 reactions per PCR) using ITS1-F as the forward primer and ITS4 as the reverse reaction. f. ITS sequences were assembled and used for Blast searches to identify the fungi recovered. Questions for Results Approximately how many animals produced at least one colony? Approximately how many lung fragments produced at least one fungal colony? What fungi were recovered? Questions for Discussion What is the significance of the fact that most lung fragments possessed at least one fungal species? Which of the fungi recovered here have been shown to produce infections in humans or non-human animals? What portion of the lungs produced colonies of Emmonsia parva (=Blastomyces parvus, = Haplosporangium parvum)? What is the significance of this?