immune variation disclosures resources and research impacts rml... · 2018-08-06 · presentation...

Immune Variation in miceResources and

Research ImpactsCory Brayton, DVM, Diplomate, ACLAM, ACVP

Director, Phenotyping CoreAssociate Professor, Molecular and Comparative Pathobiology

Johns Hopkins University, School of Medicine733 North Broadway BRB 851

Baltimore, MD [email protected]

JHU Mouse Pathobiology & Phenotyping Course Home & Links http://mcp.bs.jhmi.edu/me680712‐

phenotyping‐functional‐genetics

• No financial disclosures that I know of…

• All opinions expressed and implied in this presentation are solely those of Dr. Brayton.

• The content of the presentation does NOT represent or reflect the views of Johns Hopkins University or Johns Hopkins Health system, or of ILAR or the National Academies.

Disclosures

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Discussion Plan

1. Immune Sufficient mice Immune relevant genetic variations

2. Mission Critical Nomenclature

3. Immune deficient options

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AIMS & Conclusions

1. Expect immune variation

2. Appreciate mouse genetics and names

3. Informed selection of mice subjects

4. Understand and Use Correct nomenclature

Achieve and Report robust, relevant, reproducible and ’translatable’ research

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Your Favorites ?

1. C57BL/6

2. BALB/c

3. C3H

4. NOD

5. FVB/N

6. 129

7. OTHER?

1. Nude

2. Scid

3. NODscid

4. NSG

5. NOG

6. NRG

7. OTHER?

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Agent DZ Susceptible Intermediate Resistant

MHV ‐demyelination B6, BALB/c SJL/J

Parvo MPV1 seroconversion C3H/HeN BALB/c, DBA,

ICR B6

TMEV demyelination SJL/J, SWR, DBA/2 CBA, C3H A, B6, B10, DBA/1

Sendai Pneumonia DBA, 129 A, BALB, SWR B6, SJL

Ectromelia DBA, BALB/c, C3H, immune def B6, AKR

Mycoplasma BALB/c, C3H, A/J DBA/2, AKR B6, B10

H hepaticus A/J C3H/HeJ & NNu scid IL10‐ Rag2‐ B6, FVB/N

Lyme Arthritis BALB/c C3H B6, DBA/2

Strain Variation (infection ‘phenotypes’)

DISCLAIMER

• This is an over simplification of strain responses to these agents

• Emphasizes some responses with relatively recent data

2018 RML NIAID Brayton Immune Variation

[email protected] 2018 of 12

C57BL/6 (‘B6’)

Important/famous for:

• 1st Mouse Genome project: C57BL/6J

• ES cells for IMPC (IKMC): C57BL/6N

• Low cancer• Disease resistance i.e. the mice that didn’t die ….

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• Black, non agouti a/a• Hydrocephalus (big ventricles) • Microphthalmia• Ulcerative dermatitis • Big thymus

stress susceptible?

• Small adrenals• Acidophilic macrophage pneumonia• Amyloidosis, reactive + senile • Osteoporosis• Portal systemic shunts (J)• Cerebral vascular variations

B6 Phenotypes (a few):

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Atherosclerosis Like drugs & alcohol Dz resistance Lo tumor strain but:

Lymphoma, Histiocytic sarcoma, esp in F

C58 – thymic lymphomaC57BL/Ks ‐ hydronephrosis

B6 Genotypes (a few):Geneallele Gene , allele info System /phenotypea/a a/a Black non agoutiH2 b MHC haplotype ImmunityAhrb‐1 aryl‐hydrocarbon receptor B1 variant (immune) etcApoa2a Apolipoprotein A2 a allele Less? Senile amyloid Cdh23ahl cadherin 23 age related hearing loss 1 HearingSlc11a1s solute carrier family 11 (Nramp1) susceptible Immunity

Some Genes with variations among J and N substrainsAanat arylalkylamine N‐acetyltransferase ( lo melatonin) Endocrine/Pineal Dock2 dedicator of cyto‐kinesis 2 ImmunityNlrp12 NLR protein 12 ImmunityNnt nicotinamide nucleotide transhydrogenase deficient Gluc Metabolism

Snca Alpha synuclein 1 point mutation CNS/neuroCrb1rd8 crumbs homolog 1; retinal degeneration 8 vision9

Summarized from MGI/IMSR etc

B6 History and SUBstrains (a few)

C57BL/6NCr =

C57BL/6By C57BL/6ByJ

C57BL/6JBailey N UCSF J

adapted from http://www.envigo.com/

• Expect genotype variations

• Expect phenotype variations

• Which do you use?

• WHY?

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B6 SUBstrain options…Substrain Source

C57BL/6J C57BL/6J The Jackson Laboratory via CRL Japan, Inc. (Yokohama, Japan)C57BL/6JJcl CLEA Japan Inc. (Tokyo, Japan)C57BL/6JJmsSlc Japan SLC, Inc. (Hamamatsu, Japan)C57BL/6JEiJ The Jackson Laboratory (Bar Harbor, ME, USA)C57BL/6JOlaHsd Envigo UK, IL , NL (Harlan) (IN, USA) C57BL/6JRccHsd Envigo IL , NL (Harlan) (IN, USA) C57BL/6JBomTac Taconic (NY, USA)

C57BL/6N C57BL/6NJ The Jackson Laboratory (Bar Harbor, ME, USA)C57BL/6NCrSim Simonsen Laboratories, Inc. (Gilroy, CA, USA)C57BL/6NTac Taconic Farm Inc. ( NY, USA)C57BL/6NJcl CLEA Japan Inc. (Tokyo, Japan)C57BL/6NSeac Kyudo Co. Ltd. (Tosu, Japan)C57BL/6NCrlCrlj Charles River Laboratories Japan, Inc. (Yokohama, Japan)C57BL/6NCrl Charles River Laboratories ( MA, USA)C57BL/6NHsd Envigo MANY sites (Harlan) (IN, USA) C57BL/6NCrSlc Japan SLC, Inc. (Hamamatsu, Japan)C57BL/6ByJ The Jackson Laboratory (Bar Harbor, ME, USA)

12 Updated from Mekada et al 2015

N

B6 SUBstrain variations

B6 Substrain Source(‐/‐)

Dock2(‐/‐)

Nlrp12(‐/‐)Nnt

(‐/‐)Snca

(‐/‐)Mmrn1

(‐/‐)Rd8

C57BL/6J Jackson No YES YES No No No

C57BL/6J * Charles Riv NP NP YES No No No

C57BL/6JOlaHsd Hsd/Envigo NP NP No YES YES No

C57BL/6JRccHsd Hsd/Envigo NP NP No No No No

C57BL/6JBomTac Taconic NP NP No No No No

C57BL/6JRj Janvier NP NP NP NP NP NP

C57BL/6ByJ Jackson No NP No No No No

C57BL/6NHsd Hsd/Envigo SOME NP No No No YES

C57BL/6NRj Janvier No NP NP NP NP NP

C57BL/6NCrl Charles Riv No NP No No No YES

C57BL/6NTac Taconic No NP No No No YES

C57BL/6NCr NCI NP No NP NP NP NP

Adapted/updated from envigo.com 2015 * J mice distributed by Crl in EU NP = not published

J

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B6 SUBstrain variations

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Hypomorphic spontaneous mutations (a few):

Dock2 dedicator of cyto‐kinesis 2; altered B cell development/ migration, response to chemokines ;lack MZ B cells; Increased CD8+memory cells; Discovered in GEM on C57BL/6NHsd background. Mahajan et al 2016

Nlrp12 NLR family, pyrin domain containing 12; innate immunity, neutrophil recruitment, dendritic and myeloid cell migration

Nnt nicotinamide nucleotide transhydrogenase; encodes an integral protein of the inner mitochondrial membrane, metabolic phenotypes

Snca alpha synuclein; protein in Lewy body inclusions, implicated in Parkinson’s DzDeletion includes Mmrn1 = multimerin 1; a stored platelet and endothelial celladhesive protein.

Rd8 retinal degeneration 8; single base pair mutation in the CRB1 gene (linked to macular degeneration in humans)

Location Building Strain Dock2hsd PCR Results Indianapolis, IN 202A C57BL/6NHsd Absent Indianapolis, IN 202C C57BL/6NHsd C57BL/6BrdCrHsd‐Tyrc Absent Indianapolis, IN 217 C57BL/6NHsd B6.V‐Lepob/OlaHsd Absent Haslett, MI 206 C57BL/6NHsd Present repopulated 2017/8Frederick, MD 208 C57BL/6NHsd Present repopulated 2017/8Houston, TX* 211 C57BL/6NHsd Absent Dublin, VA 231 C57BL/6NHsd CB6F1/Hsd B6C3F1/Hsd B6D2F1/Hsd Present repopulated 2017/8Livermore, CA 237 C57BL/6NHsd Absent Mexico City, Mexico 650 C57BL/6NHsd Present repopulated 2017/8Gannat, France Isolators C57BL/6JOlaHsd C57BL/6JRccHsd B6.V‐Lepob/OlaHsd Absent Horst, Netherlands Barrier 2 C57BL/6JOlaHsd C57BL/6JRccHsd Absent

Horst, Netherlands Barrier 2 C57BL/6NHsd Present Rehovot, Israel 640 C57BL/6JOlaHsd C57BL/6JRccHsd Absent Jerusalem, Israel 610 C57BL/6JOlaHsd Absent Udine, Italy 700W C57BL/6JOlaHsd, C57BL/6JRccHsd Absent Udine, Italy 700E C57BL/6JOlaHsd Absent Blackthorn, England C Block B6.V‐Lepob/OlaHsd Absent Bresso, Italy Isolators C57BL/6JOlaHsd C57BL/6JRccHsd Absent Bresso, Italy Isolators C57BL/6NHsd Present repopulated 2017/8Seoul, Korea N/A C57BL/6NHsd Results pending Borchen, Germany* Winkelmann C57BL/6NHsd Results pending Tests were conducted on foundation colony breeding cages. Genetic testing was performed at the Envigo Genetic Testing laboratories in Piscataway, NJ using qPCR technology. *Facility closed prior to 2016

Dock2hsd in C57BL/6NHsd 6/9/16

Mutation eliminated from colonies by restock with ‘original’ Indianapolis stock…. What about GEM backcrossed to C57BL/6NHsd ?

Arose >20y ago…15

Useful B6 references • Zurita et al. 2010. Genetic polymorphisms among C57BL/6 mouse inbred strains. Transgenic Res.

• Simon & al. 2013. A comparative phenotypic and genomic analysis of C57BL/6J andC57BL/6Nmouse strains.

• Kraev. 2014. Parallel universes of Black Six biology.• Mekada et al. 2015. Development of SNP markers for C57BL/6N‐derived mouse inbred strains.

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• Abolins et al. 2017. The comparative immunology of wild and laboratory mice, Mus musculus domesticus. Nature. B6 vs wild mice (& humans) …

• Beura et al. 2016. Normalizing the environment recapitulates adult human immune traits in laboratory mice. Nature. B6 vs pet store mice (& humans)….

• Seok et al. 2013. Genomic responses in mouse models poorly mimic human inflammatory diseases. PNAS. Takao & Miyakawa. 2015. Genomic responses in mouse models greatly mimic human inflammatory diseases. PNAS. (Same Data)

B6 representing all mice vs human…

B6 represent ALLLaboratory mice ?

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WILDA, BALB, C3

DBA

129

C57, C58Group 1: Bagg albino derivatives

Group 2: Swiss

Group 3: Japanese & New Zealand inbred

Group 4: Abbie Lathrop’s C57/58

Group 5: Castle's 129 etc

Group 6: CC Little's DBA & related

Group 7: wild‐derived

Which Mice ?Could represent all Lab mice??

Petkov et al 2004. An efficient SNP system for mouse genome scanning elucidating strain relationships. Genome Res. 2004 Sep;14(9):1806‐11.

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NEXT:

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BALB/c…..• Famous For Monoclonal Antibodies Used to study Infectious Disease

• Likely Killers (CoD = Cause of Death) Each other – esp males Heart disease, diverse tumors

• Also see Acallosity; Cardiac calcinosis, Thrombi, Cardiomyopathy; Vaginal septa, imperforate vagina, poor fertility

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BALB/c Phenotypes (a few):

Cardiac Conditions

• Epicardial mineralization Right Ventricle (RVFW)

• Thrombi left atrium

• ‘Cardiomyopathy’ Inflammation Degeneration Fibrosis

Miscellaneous Tumors• Plasma cell myeloma Monoclonal AB

• Rhabdomyosarcoma• Salivary Myoepithelioma

• Mammary • Harderian gland• Adrenal cortical • Thyroid

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1937 1937/81938/9

1913

1947

BALB/c History & SUBstrains(a few): Family tree (Pedigree) of Bagg’s albino mice

• Which do you use?

• Why?

Andervont

N strains

ScottJ

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1950

NCI Pt

BALB/c substrain options

Andervont

BALB/cAn BALB/cAnBy Bailey

o BALB/cByJ

BALB/cAnN NIHo BALB/cAnNCrlo BALB/cNHsdo BALB/cAnNTac

BALB/cAnHe Heston Etc… Pt, Ski, Arg,

• Scott J

• BALB/cJ BALB/cJBomTac BALB/cOlaHsd

♦ More – BUT not so available, e.g. – BALB/cWtEiJ

Separated c 1932 ~F26

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Female Biasin Infectious Dz Research?• ….Scott did not select or purposely breed BALB/c mice for aggressive behavior ….In 1942 he notes that weanling BALB/c males could be housed 5 to a cage ... Further BALB/c albinos became lethargic in warm weather.

• …These observations put us at a loss to explain why BALB/cJ males are viciously aggressive when compared with their docile BALB/cAn ….

From Potter 1985. History of the BALB/c Family• https://link.springer.com/chapter/10.1007/978‐3‐642‐70740‐7_1

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BALB/c genotypes (a few):Strains Geneallele Gene , allele info System or phenotype

BALB/cByJBALB/cJ

c/c A/A Tyrp1b/Tyrp1b Tyrc/Tyrc albino

H2 d MHC haplotype immune

Hld hippocampal lamination defect Brain

Apoa2b Apolipoprotein A2 – b variant (Hdlq5) Hier HDL (lo ASSAM)

BALB/cByJBALB/cAnEtc?

Prkdcbalb/cprotein kinase, DNA activated, catalytic polypeptide (Balb/c allele has less effect than scid)

Immunity ….radiosensitivity etc

BALB/cByJ Acadsdel‐J acyl‐Coenzyme A dehydrogenase deficiency organic aciduria

BALB/cByJ Ahrb‐2 aryl‐hydrocarbon receptor B2 variant Immunity

BALB/cByJ Cdh23ahl cadherin 23 (otocadherin) age related hearing loss 1 Hearing

BALB/cByJ Mdmg1 mandibular morphogenesis 1 longer dorsal edge Skeletal

From http://www.findmice.org/IMSRSearchForm.jsp etc27



Another example…

• …gene mutation causing complete unresponsiveness [to LPS] occurred after 1960 and by 1968 was inbred into the C3H/HeJ colony… Glode & Rosenstreich 1976.

https://www.ncbi.nlm.nih.gov/pubmed/792337

C3H SUBstrains…. & the story of Tlr4lps

C3H/StC3H/CrC3H/BiC3H/HeC3H/HeNC3H/HeJC3H/HeBFeJC3H/DiSnC3H/AvyC3H/Bts

Fire Jax

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Tlr4lps‐del

In C57BL/10ScNJ

Formerly known as: C57BL/10ScCr ‐ Poltorak et al. (1998). C57BL/10Cr ‐ Coutinho & Meo (1978).

Another Spontaneous Tlr4mutation

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• Famous for ES cells TGCT – testicular teratomas (Ter etc genes )

• Likely/possible killers (CoD = Cause of Death) Acidophilic macrophage pneumonia Megaesophagus, arteritis, tumors

• Also Substrain variation…. Acallosity Deaf ? Otitis ?

129

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ES cells & who they came from:

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ES Cell Line Strain

AB1 129S7/SvEvBrd-Hprt+

AB2.1 129S7/SvEvBrd-Hprtb-m2

AB2.2 129S7/SvEvBrd-Hprtb-m2

AK7 129S4/SvJaeSor

BALB/c-I BALB/Ci

BK4 129P2/OlaHsd

BL/6-III C57BL/6

Bruce 4 C57BL/6

Bruce 7 C57BL/6

C1 129X1/SvJ

C13 129X1/SvJ

C57BL/6 C57BL/6NTac

C57BL/6Hprt C57BL/6J-Hprtb-m3

CB1-4 C57BL/6J x (Rb(11.16)2H x Rb(16.17)32Lub)F1

CBA CBA/CaOlaHsd

CC1.2 129S7/SvEvBrd

CGR8 129P2/Ola

CJ7 129S1/Sv-p+ Tyr+ Kitl+

CK35 129S2/SvPas

CMTI-1 129/Sv

CMTI-2 C57BL/6

CP-1 129S6/SvEvTac

CT129 129/Sv

D3 129S2/SvPas

D3a2 129S2/SvPas

D3H 129S2/SvPas

DBA-252 DBA/1LacJ

E14 129P2/OlaHsd

E14.1 129P2/OlaHsd

E14K 129P2/OlaHsd

E14TG2a 129P2/OlaHsd

EK.CCE 129S6/SvEvTac

ENS 129/Sv

ESF 116 CBA

ESF 122 CBA

ESF 48/1 129P2/Ola

ESF 55 129P2/Ola

ESF 58/2 129P2/Ola

GK129 129P2/OlaHsd

GS1 129/Sv

GSI-1 129X1/SvJ

GSIB-1 C57BL/6

H1 129/Sv

HM-1 129P2/OlaHsd

IB10 129P2/OlaHsd

J1 129S4/SvJae

KG1/KG-1 129S6/SvEvTac

Lex-1 129S5/SvEvBrd

LSW1 129X1/SvJ

mEMS32 129P3/JEmsJ

MESC 20 129P2/OlaHsd

MPI 65-3 C57BL/6J-Hprtb-m3

MPI-12D 129S6/SvEvTac

MPI-12G 129S6/SvEvTac

MPI-17A 129S6/SvEvTac

MPI-17E 129S6/SvEvTac

MPI-48.1 C57BL/6J-Hprtb-m3

MPI-71.6 C57BL/6J-Hprtb-m3

MPI-II 129/Sv

MPI53.1 C57BL/6J-Hprtb-m3

MPI76.11 C57BL/6J-Hprtb-m3

MRL-+/+ 3 MRL/MpJ

P1 129S2/SvPas

Pat5 129X1/SvJ

PC3 129S4/SvJae

PJ1-5 129X1/SvJ

PJ5 129X1/SvJ

R1 (129X1/SvJ x 129S1/Sv)F1-Kitl+

REK1 129X1/SvJ

REK2 129X1/SvJ

REK3 129X1/SvJ

REK4 129X1/SvJ

RF8 129S4/SvJae

RW-4 129X1/SvJ

SM1 129S6/SvEvTac

TC 129S6/SvEvTac

TC-1 129S6/SvEvTac

TG3 129S6/SvEvTac

TG4 129S6/SvEvTac

TG6 129S6/SvEvTac

TL1 129S6/SvEvTac

TT2 (C57BL/6 x CBA)F1

v17.2 (BALB/cJ x 129S4/SvJae)F1

v6.4 (C57BL/6J x 129S4/SvJae)F1

W12 129S6/SvEvTac

W2 129S6/SvEvTac

W3 129S6/SvEvTac

W4 129S6/SvEvTac

W5 129S6/SvEvTac

W9.5 129S1/Sv-p+ Tyr+ Kitl+

WB6a C57BL/6

WB6b C57BL/6

WB6d C57BL/6

WW6 STOCK 129/Sv and C57BL/6J and SJL

NOW: Search IMSR http://www.findmice.org/index.jsp for parental strains of ESC lines

USE THIS SITE:http://www.informatics.jax.org/mgihome/nomen/index.shtml

16 129 'strains’… 13 = ‘129Sv’…

http://www.informatics.jax.org/mgihome/nomen/strain_129.shtml

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Abbreviated designation Full designation Former designation Coat Color Coat Color Genotype

129P1-Lama2dy 129P1/Re-Lama2dy 129/Re-Lama2dy pink-eyed chinchilla Aw/Aw p Tyrc-ch /p Tyrc-

chLama2dy /Lama2dy

129P1 129P1/ReJ 129/ReJ pink-eyed chinchilla Aw/Aw p Tyrc-ch/p Tyrc-ch

129P2 129P2/OlaHsd 129/OlaHsd pink-eyed chinchilla Aw/Aw

p Tyrc-ch/p Tyrc-ch

129P3 129P3/J 129/J pink-eyed light chinchilla or albino

Aw/Aw p Tyrc-ch/p Tyrc

Aw/Aw p Tyrc/p Tyrc

129X1 129X1/SvJ 129/SvJ pink-eyed light chinchilla or albino

Aw/Aw p Tyrc-ch/p Tyrc

Aw/Aw p Tyrc/p Tyrc

129S1 129S1/Sv-p+Tyr+KitlSl-J/+ 129/Sv-p+Tyr+KitlSl-J/+ white-bellied agouti Aw/Aw

129S1 129S1/SvImJ 129/Sv-p+Tyr+Kitl+/J white-bellied agouti Aw/Aw

129S2 129S2/SvPasCrl 129/SvPas white-bellied agouti Aw/Aw

129S4 129S4/SvJae 129/SvJae white-bellied agouti Aw/Aw

129S5 129S5/SvEvBrd 129/SvEvBrd white-bellied agouti Aw/Aw

129S6 129S6/SvEvTac 129/SvEvTac white-bellied agouti Aw/Aw

129S7 129S7/SvEvBrd-Hprtb-m2 129/SvEvBrd-Hprtb-m2 white-bellied agouti Aw/Aw

129S8 129S8/SvEv-Gpi1c Hprtb-m2@J 129/SvEv-Gpi1c Hprtb-m2@J white-bellied agouti Aw/Aw

129T1 129T1/Sv-p+ Tyrc-ch Ter/+@Na 129/Sv-p+ Tyrc-ch

Ter/+@Nawhite-bellied agouti chinchilla Aw/Aw Tyrc-ch/Tyrc-ch

129T2 129T2/SvEms 129/SvEms-Ter+? white-bellied agouti chinchilla Aw/Aw Tyrc-ch/Tyrc-ch

129T2 129T2/SvEmsJ 129/SvEms-Ter+?/J white-bellied agouti chinchilla Aw/Aw Tyrc-ch/Tyrc-

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129____ whatever…http://www.informatics.jax.org/mgihome/nomen/strain_129.shtml

‘P’ indicates origin from Parental stock; ALL of the rest are/were 129Sv…..

‘S’ indicates origin from a line that had carried Sl (Steel allele on the Kitl gene, homozygous lethal);

‘T’ indicates that the line carries/d Ter (teratoma) gene;

‘X’ indicates that the line was genetically contaminated. (Festing et al. 1999; Simpson et al. 1997)

129X1/SvJ, has contributions by

o C57BL/6J on Chromosomes 5, 7, 14, 18, 19, also

o BALB/cJ on Chromosomes 7, 8, 10, 18, 19, X,

o suggesting CBF1 hybrid as contaminant. (Petkov & al. 2004)http://www.informatics.jax.org/mgihome/nomen/index.shtml



129 genotypes (a few):

Strains Geneallele Phenotypes129 A, w, p, d, sl Colors

129 H2b BUT may differ at minor loci graft rejection129P3J Ahrd aryl‐hydrocarbon receptor allele d 129P1/ReJ 129P3J129X1J Cdh23ahl cadherin 23 (otocadherin) age related hearing loss 1

129P1/ReJ 129P3J 129P4 129T2 129S1129X1J

Disc1del Disrupted in schizophrenia

129P3J Rmcfr resistance to MCF virus

129 @GSF Pde6brd1 Blind ‐ Retinal degeneration ‐ Dalke & al 2004

129S2PasCrl Slc3a1m1Crl Cystinuria UROLITHIASIS

From http://www.findmice.org/IMSRSearchForm.jsp etc34

• Important/ Famous for Prominent pronuclei (transgenesis) Fecund (nice mice w big litters)

• Likely/possible killers (CoD = Cause of Death) Seizures ? Maybe tumors

• Also Mammary hyperplasia, EMT w Pituitary tumors; Lung tumors, Skin sarcomas

• Big, blind, albino mice big litters, not so likely to kill each other

FVB/N

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Outbred Swiss origin

BLIND rd/rd1

FVB/N genotypes (a few):

Strains Geneallele Gene , allele info System or phenotypeFVB/NJ c/c A/A Tyrc/Tyrc Albino

H2q MHC haplotype

Ahrb‐2 aryl‐hydrocarbon receptor B2 variant Immunity

Fv1 Friend virus susceptibility 1 Immunity

Hc0 C5, hemolytic complement deficient immunity

Pde6brd1 Retinal degeneration 1 phosphodiesterase 6B, cGMP, rod receptor, beta polypeptide Vision

Apoa2b Apolipoprotein A2 – b variant (Hdlq5) Hier HDL

Disc1del disrupted in schizophrenia 1 Neuro??

From http://www.findmice.org/IMSRSearchForm.jsphttps://www.jax.org/strain/001800 etc36

STRAIN MATTERS.Phenotypes in ‘Normal’ ‘wt’ (a few):

129? Teratomas (Ter), lung tumors, acallosity, AMP, …

A/J Lung tumors, anomalies, amyloid, muscular dystrophy

AKR Thymic Lymphoma…

BALB/c Plasmacytoma etc tumors, heart dz, acallosity, kill each other

C3H TUMORS ‐ Mammary, Liver

C57BL/6 Microphthalmia, Hydrocephalus , MUD, Osteoporosis, Presbyacusis, Amyloid, AMP,

DBA Deaf, seizures, glaucoma, autoimmune

FVB/N? Blind, seizures, mammary/pituitary dz

NOD Diabetes, immunoweird

SJL/J Lymphoma, muscular dystrophy, kill each other

DEAF C57BL/6, BALB, DBA, etc

BLIND rd1 C3H, CBA, SJL, SWR, FVB 37

BLIND rd/rd1

Immune Relevant Genotypes (a few):STRAIN MHC c5 Ahr ApoA2 Il3ra Rmcf Sclc11

(Nramp) OTHER

129 H2 b ~~~~~~ Beware of substrain variations ~~~ Slc11a1R

AA/J H2 a Hc0 Ahrb‐2 Apoa2c Il3ram1 Rmcfs Tnfrsf13c Bcmd1‐A

AKR H2 k Hc0 Ahrb‐2 Apoa2a Il3ram1 Rmcfs

BALB/cBALB/cBy H2 d Ahrb‐2 Apoa2b Slc11a1s Prkdcbalb/c

C3HC3H/HeJ H2b‐2 Ahrb‐2 Apoa2b Slc11a1R Tlr4Lps‐d

C57BL/6JC57BL/6N H2 b Ahrb‐1 Apoa2a Slc11a1s Nlrp12B6J

Dock2Hsd

DBA/1DBA/2

H2 qH2 d Hc0 Ahrd Rmcfr Slc11a1s

Slc11a1R

FVB/N H2 q Hc0 Ahrb‐2 Apoa2b

‘MRLlpr’ H2 k Il2m1 Foxq1sa‐J Faslpr

NOD H2 g7 Hc0 Il2m1 Sirpa

SJL/J H2s Ahrd Apoa2c Rmcfs Il2m1

Ceacam1Hv2‐r39

Immune Variations, Strain Associated(More) Innate (More) Adaptive/Acquired

TH1 bias e.g B6 TH2 bias e.g. BALB/cHc c5 hemolytic complement MHCSclc11a1 solute carrier family 11a member 1

(formerly Nramp1) Tlr4 Toll like receptor 4

Natural Killer Cell function ‐ Variation in NK complex Klra (Ly49) Klrb (Nkrp) etc

Il2m1 Il 2; mutation 1 = Hypo‐active variant of IL‐2 with decreased T cell activation

NAIP neuronal Apoptosis inhibitor proteins;Nlrp‐ NOD like receptor protein polymorphisms

Tcrbv8 T cell receptor beta variable 8

Sirpα in NODphagocytosis Slamf signaling lymphocyte activation molecule family (CD48) polymorphisms

Cathepsin E function Dock2 ‐ disruptor cytokinesis 2Mx1, Mx2myxovirus (influenza) resistance 1,2

strain marker

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SEX MATTERS too.

• Prevalence of sexual dimorphism in mammalian phenotypic traits. Nat Commun. 2017 Jun 26;8:15475. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5490203/ How many authors does it take to convince a scientist?

• Karp et al: Mason J, Beaudet AL, BenjaminiY, Bower L, Braun RE, Brown SDM, Chesler EJ, Dickinson ME, FlennikenAM, Fuchs H, Angelis MH, Gao X, Guo S, Greenaway S, Heller R, HeraultY, Justice MJ, Kurbatova N, Lelliott CJ, Lloyd KCK, Mallon AM, Mank JE, Masuya H, McKerlie C, Meehan TF, Mott RF, Murray SA, Parkinson H, Ramirez‐Solis R, Santos L, Seavitt JR, SmedleyD, SorgT, Speak AO, Steel KP, Svenson KL; International Mouse Phenotyping Consortium, Wakana S, West D, Wells S, Westerberg H, Yaacoby S, White JK. +159 Collaborators: bataY, Suzuki T, Tamura M, Kaneda H, FuruseT, Kobayashi K, Miura I, Yamada I, Tanaka N, Yoshiki A, Ayabe S, Clary DA, Tolentino HA, Schuchbauer MA, Tolentino T, Aprile JA, Pedroia SM, Kelsey L, Vukobradovic I, Berberovic Z, Owen C, Qu D, Guo R, Newbigging S, Morikawa L, Law N, Shang X, Feugas P, Wang Y, Eskandarian M, Zhu Y, Nutter LMJ, Penton P, LaurinV, Clarke S, Lan Q, Sohel K, Miller D, Clark G, Hunter J, Cabezas J, BubshaitM, Carroll T, Tondat S, MacMaster S, Pereira M, Gertsenstein M, DanismentO, Jacob E, Creighton A, Sleep G, Clark J, Teboul L, Fray M, Caulder A, Loeffler J, CodnerG, Cleak J, Johnson S, Szoke‐Kovacs Z, RadageA, Maritati M, Mianne J, Gardiner W, Allen S, Cater H, Stewart M, Keskivali‐Bond P, Sinclair C, Brown E, Doe B, Wardle‐Jones H, Grau E, Griggs N, Woods M, Kundi H, Griffiths MND, KippC, DT, VancollieVE, Pearson SA, Gates AS, Sanderson M, Shannon C, Anthony LFE, Sumowski MT, McLaren RSB, SwiatkowskaA, Isherwood CM, Cambridge EL, Wilson HM, Caetano SS, MazzeoCI, DabrowskaMH, LillistoneC, Estabel J, Maguire AKB, Roberson LA, PavlovicG, Birling MC, Marie WD, Jacquot S, Ayadi A, Ali‐Hadji D, Charles P, André P, Le Marchand E, El AmriA, Vasseur L, Aguilar‐Pimentel A, Becker L, Treise I, Moreth K, StoegerT, AmarieOV, Neff F, WurstW, Bekeredjian R, Ollert M, Klopstock T, Calzada‐Wack J, Marschall S, Brommage R, Steinkamp R, Lengger C, Östereicher MA, Maier H, Stoeger C, Leuchtenberger S, YildrimA, Garrett L, Hölter SM, ZimprichA, Seisenberger C, Bürger A, Graw J, EickelbergO, Zimmer A, Wolf E, Busch DH, Klingenspor M, Schmidt‐Weber C, Gailus‐DurnerV, Beckers J, Rathkolb B, RozmanJ. Melvin DG, Raj NPS, Holroyd SA, Gannon DJ, Alcantara R, Galli A, Hooks YE, Tudor CL, Green AL, Kussy FL, Tuck EJ, SiragherEJ, Maguire SA, Lafont41

SEX MATTERS too.

AUTOIMMUNE• Graves• Hashimoto• RA• SLE• T1DM

INFECTIOUS

• HIV• Flu• Toxo• Legionella• Malaria• Zika

42

NON repro CA • Bladder• Bowel• Kidney • Leukemia• Liver• Lung• Melanoma• Esophagus• Stomach

• Ebola• MERS• HepB• Campy• Schistosomiasis• Amebiasis• Aspergillosis

Adapted f rom Chervonsky 2018Klein & Flanagan 2016

• Correct strain names identify strain(s) of origin, and where it has been, and came from

• Correct mutant names tell a lot about the mice & mutations

• Incorrect names compromise communication, reproducibility, translational research

Nomenclature: mission critical in scientific communication…

USE CORRECT NAMES. REQUIRE CORRECT NAMES when reviewing & editing. Get a Lab Code if you make or breed mice Its FREE $ & EASY! USE THIS SITE:http://www.informatics.jax.org/mgihome/nomen/index.shtml43

Strain First - Genotype/Laboratory codes

• B6.129P2-Apoa1tm1Unc/J

• B6.I29P2-ApoaItmIUnc/J

• B6;129X1-Apoa5tm1Lap Apoc3tm1Lap

• B6;I29XI-Apoa5tmILap Apoc3tmILap

• B6;CBA-Tg(APOC1)1Bres/J

• B6;CBA-Tg(APOCI)IBres/J

Inbred Nomenclature

44

Background strain(s) ?Congenic ? Mutation strategy ?Inserted gene(s) ?Disrupted gene(s)? Who made it? Where is it available ?

USE THIS SITE (+TUTORIAL):http://www.informatics.jax.org/mgihome/nomen/index.shtml

Outbred Nomenclature

Lab code1st: STOCK name(etc info) e.g.

CRL:CD-1®(ICR)BRCRL:CFW®(SW) BR CRL:CF-1® BR (NOT Swiss)Cr:NGP(S)Cr:NIH(S)

45

Hsd:ICR(CD-1®)Hsd:ND4Hsd:NIHS IcrTac:ICR Tac:SWBomTac:NMRI

Lab Code indicates Source BR = barrier reared; cc = closed colony

USE THIS SITE (+TUTORIAL):http://www.informatics.jax.org/mgihome/nomen/index.shtml

• ‘genetically undefined’ vs ‘outbred’

NOT inbred but also Famous..

abbrev origin

CD1 Swiss

ICR Swiss

ND4 Swiss

NMRI Swiss

Swiss Swiss

SW (Swiss Webster) Swiss

[CC Collaborative Cross (8 strains ) RI strains]

DO CC RI (Recombinant Inbreds)

4WC, Het Other heterogeneous stocks

46



INcompleteness of background strain reporting in recent publications

Background strain notation 2010 2011 2012 2013 2014 Combined totals or averages

Completely addressed 54 56 49 52 43 Combined total: 254

Incompletely addressed 77 64 55 86 80 Combined total: 362

Incomplete due to C57BL/6 43 43 40 52 48 Combined total: 226

Total number of articles 126 123 105 138 124 Combined total: 616

% incomplete total 61 52 52 62 64.5 Average: 58.5% of incompletes due to C57BL/6 56 67 73 60.5 60 Average: 63

Fontaine & Davis 2016. Diabetes. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4686949 /

Mice/Names

•CONCLUSIONS: Strain matters. Sex matters. Beware of incomplete or inaccurate names… Be aware of nomenclature …

Final Exam : (Your research! … and reporting it …)oWHY do you use a specific strain or substrain?oRelevant phenotypes/genotypes to your research?oWould other strains be useful to your research? oDo you use both sexes? & Analyze data separately? 48

Your Favorite Immune Deficient Mice?

1. Nude

2. Scid

3. NODscid

4. NSG

5. NOG

6. NRG

7. OTHER?

• Inbred? BALB/c C.B17 B6 C3H NOD Other?

• Non inbred? (genetically undefined, Usually Swiss ‘outbred’??)

• Not sure?49

Making ‘better’ immunodeficient mice

• Better for xenografts• Better for humanizing: replacing mouse immune (or other) systems with human.

• Less early death dt thymic lymphoma

• Valkenburg, Williams 2011

53

Lymphomas ??

Lymphomas

Lymphomas

Sirpa & cd47

Background Strain Effects (J) Back ground

InnateNK etc Leaky H2 NAME Characteristics (Phenotypes)

BALB/cByJ Normal Yes T d CByJ.Cg‐Foxn1nu/JNormal B cells etc; RadioSensitive: (Prkdcbalb/c ) Extra thymic T cells with agePOOR breeders, female fertility nu/+ F x nu/nu M

Normal High d CBySmn.CB17‐Prkdcscid/JNO functional B and T cells; RadioSensitive: (Prkdcbalb/c ) HIGH NK, complement activity (normal APC function)Thymic lymphomas but < NOD.CB17‐Prkdcscid/SzJ

C57BL/6J Normal NO b B6.129S7‐Rag1tm1Mom/JNO functional B and T cellsHIGH NK activity (normal APC, complement)

Normal High b B6.CB17‐Prkdcscid/SzJNO functional B and T cellsHIGH NK, complement activity (normal APC function)

NOD/LtSzJ Low NO g7 NOD.129S7(B6)‐Rag1tm1Mom/JNO functional B and T cellsPre‐B cell > Thymic lymphomas ( 10.5mo)Somewhat RadioResistant

Low NO g7 NOD.Cg‐Rag1tm1Mom Prf1tm1Sdz/SzNO functional B &T cells; NO NK cell activity; RadioResistant (to 8gy): Thymic lymphomas (short lifespan ~8.5mo)

Low Low g7 NOD.CB17‐Prkdcscid/SzJ

NO functional B /T; NOD background low NK activity, NO hemolytic complement, defects in myeloid development, poor APC functions ‐‐RadioSensitive: tolerates up to 4 Gy; Sirpa

Thymic lymphomas (short lifespan ~8.5mo)

Low Low g7 NOD.Cg‐Prkdcscid B2mtm1Unc/JNO functional B and T cellsNo MHC 1 expression ~NO NK activity Thymic lymphomas (short lifespan ~6.3mo); Hemachromatosis dt?

Low NO g7 NOD.Cg‐Prkdcscid Il2rgtm1Wjl/SzJNO functional B and T cells; NO NK cell activityLymphoma‐resistant and long‐lived > 16m

NIH stock Normal Yes T q NU/J (Foxn1nu) NIH outbred nude stock inbred at TJL54

adapted from Jax

notes #501, 2006

Some NOD/shiLtJ genotypes

Strains Geneallele Gene , allele info System or phenotype

NOD/shiLtJ c/c A/A Tyrc/Tyrc Albino

H2g7 MHC haplotype Kd, Aad, Abg7, Enull, Db Immunity

Cdh23ahl1 Otocadherin age related hearing loss 1 hearing

Hc0 C5, hemolytic complement deficient immunity

Il2m1 Interleukin2 hypoactive polymorphism Immunity

Sirpa

Signal regulatory protein (SIRP) locus variant encodes receptors that mediates activating &inhibitory signals;associated with type 1 diabetes (T1D); NOD variant hi affinity binding Hu Cd47 ‐> hier hu graft success

immunity

57 From MGI, IMSR http://www.findmice.org/IMSRSearchForm.jsp etc



B6 v BALB/c v NODimmunity…

B6 BALB/c NODH2 b H2 d H2 G7

TH1 bias IL12 TH2 bias IL4More cell mediated More humoral responseInt‐Hi NK more Ly49Strain Specif Klra NK complex

Int‐ less NK Strain Specif Klra NK complex

Lo NK Strain Specif Klra NK complex

More macrophage activity, TNFa, IL12, bacterial killing; more local/less systemic response

Less macrophage activity, TNFa, IL12, bacterial killing; more systemic & acute phase response,

DC/Mac effectsSirpa variation Hu Cd47 affinityHc0 effects on MAC etc

B6J / B6N variation in Th17 (microbiome?)

Prkdcbalb/c radiosensitivityDefective repair DS DNA

58

More Foxn1nu optionsSource Background Name INbred AKACrl BALB/cAnN CAnN.Cg‐Foxn1nu/Crl + BALB/c Nude

CD1 (S) Crl:CD1‐Foxn1nu CD1 nude

NIH Nu Crl:NU‐Foxn1nu Nu/nu NIH nude

Hsd Ola NIH Nu Hsd:Athymic NudeFoxn1nu NIH nude

Envigo NMRI (S) HsdCpb:NMR‐Foxn1nu Swiss Nude

CD1 (S) HsdHli:CD1‐Foxn1nu CD1 nude

ICR (S) HsdOla:ICRF‐Foxn1nu Swiss Nude

MF1 HsdOla:MF1‐Foxn1nu MF1 nude

BALB/cOla BALB/OlaHsd‐Foxn1nu + BALB nude

Tac Bom B6N/Tac B6.Cg/NTac‐Foxn1nu + B6 nude

BALB/cAnN C.Cg/AnNTac‐Foxn1nu + BALB/c nude

BALB/cBom C.Cg/AnBomTac‐Foxn1nu + BALB/cA nude

(BALB x) NIH(S) CrTac:NCr‐Foxn1nu NIH Nude

NMRI (S) BomTac:NMRI‐Foxn1nu Swiss nude

NIH(S) NTac:NIHS‐Foxn1nu Swiss Nude59

IF the name begins with the SOURCE, it is OUTbred

(or not inbred)

May not be as

homogeneous as inbred BUT May be more robust & easier to breed /

maintain….

Which are Swiss?

More Prkdcscid optionsSource Back

ground Name INbred AKA

Crl C.B17 CB17/Icr‐Prkdcscid/IcrCrl + CB17 scid

SHO Crl:SHO‐PrkdcscidHrhr Scid hairless

Hsd Ola BALB/cJ BALB/cJHan™Hsd‐Prkdcscid + BALB/c Scid

Envigo C.B‐17 C.B‐17/IcrHsd‐Prkdcscid + CB17 scid

C3H C3H.C‐Prkdcscid/IcrSmnHsd + C3H scid

ICR (S) HsdIcr:Ha(ICR)‐Prkdcscid Scid

NOD NOD.CB17/JHliHsd‐Prkdcscid + NODscid

Tac Bom C.B17 C.B‐Igh‐1b/IcrTac‐Prkdcscid + CB17 scid

IcrTac IcrTac:ICR‐Prkdcscid Scid ‐ Swiss

C.B‐17 C.B‐Igh‐1b/GbmsTac‐Prkdcscid‐Lystbg + scid‐beige

NOD + NOD/MrkBomTac‐Prkdcscid + NODscid

NOD NOD.Cg‐Prkdcscid Il2rgtm1Sug/JicTac + CIEA NOG60

IF the name begins with

the SOURCE, it is OUTbred (or not inbred)

May not be as homogeneous as inbred BUT May be more robust & easier to breed / maintain

More options

NSG or NOG?• NOD.Cg‐Prkdcscid Il2rgtm1Wjl/SzJ

NSG ‐ Il2rg null – Schultz at J ‐ (Shultz & al. 2005).

• NOD.Cg‐Prkdcscid Il2rgtm1Sug/JicTac NOG ‐ Il2rg deficient – CIEA (Jic) – Japan (Ohbo & al. 1996

Source Background Name INbred AKACrl C.B‐17 CB17.Cg‐PrkdcscidLyst bg/Crl + Scid beige

NIH(S) Crl:NIH‐Lyst bg Foxn1nu Btk xid NIHIII nude

HSD Ola CBA CBA/HNOlaHsd‐Btkxid + XID

Envigo NIH(S) Hsd:NIHS‐Lyst bgFoxn1nuBtkxid NIHIII nude

B6 C57BL/6OlaHsd‐Lystbg + BeigeC.B‐17 C.B‐17/IcrHsd‐PrkdcscidLystbg + Scid beige

Tac C.B17 C.B‐Igh‐1b/GbmsTac‐Prkdcscid‐Lystbg + Scid beige

NIH(S) Tac:NIHS‐Lystbg Foxn1nu Btkxid NIHIII nude

NOD NOD.Cg‐Prkdcscid Il2rgtm1Sug/JicTac + CIEA NOG

BALB/c C.Cg‐Rag2tm1Fwa Il2rgtm1Sug/JicTac + CIEA BRG

61

62

Immunodeficient options CONCLUSION: LOTS OF OPTIONS.

Final Exam:oDo you know WHY you use a specific ID mouse?oDo you order from the same vendor ? oDo you order exactly the same mice ? oDo you check the health reports?oDo you order from the same room/isolator?oEvery time? oDO they stay clean in your facility ?oAre You Sure?

63



AIMS & Conclusions

1. Expect immune variation

2. Appreciate mouse genetics and names

3. Informed selection of mice subjects

4. Understand and Use Correct nomenclature

Achieve and Report robust, relevant, reproducible and ’translatable’ research

64

WILDA, BALB, C3

DBA

129

C57, C58

Who should represent ALL mice (& humans)??Group 1: Bagg albino derivatives

Group 2: Swiss

Group 3: Japanese & New Zealand inbred

Group 4: Abbie Lathrop’s C57/58

Group 5: Castle's 129 etc

Group 6: CC Little's DBA & related

Group 7: wild‐derived65Petkov et al 2004. An efficient SNP system for mouse genome

scanning elucidating strain relationships. Genome Res. 2004

Headlines:

•We CAN do better.

https://www.the‐scientist.com

66

Thanks!! • Hosts !!• Audience !!• Mice & GEM

• Nadine Forbes

• MCP & core faculty

• LAM & Vet path trainees NIH T32 RR0077022

That’s all folks!

67

68

Mighty Mouse

Jargon

• SPF = Specific Pathogen Free Defined by the Exclusion list… (= tested/excluded agents)

• Gnotobiotic = defined flora ASF = altered Schaedler’s flora

• Axenic = germ free• Autochthonous flora (indigenous flora) Microbiome/Microbiota

• Allochthonous flora (transient flora).

69



• Find Gene info & correct/current names (MGI) http://www.informatics.jax.org/genes.shtml http://www.informatics.jax.org/mgihome/nomen/index.shtml (with nomenclature tutorial)

• More on strains/phenotypes https://phenome.jax.org/projects/Brayton1

• Mouse Pathobiology & Phenotyping Course & Lab manual (free to download) http://mcp.bs.jhmi.edu/me680712‐phenotyping‐functional‐genetics

RESOURCES (Mice)

70

RESOURCES: Find GEM(Genetically Engineered Mice) • IMSR – Induced Mutant Strain Resource Database find mice in multiple resources http://www.findmice.org/index.jsp

• IKMC – International Knockout Mouse Consortium (and repositories) http://www.mousephenotype.org/ (IMPC)

• EMMA – European Mutant Mouse Archive• MMRRC – Mutant Mouse Research Resource Centers (c 1998)

• RIKEN ‐ Japan

71

• Mouse Phenome Database (MPD) Emphasizing J strains; protocol detail; published studies; http://phenome.jax.org/

• EUROPHENOME GEM – IMPRESS (EMPRESS) protocols as applied in EUMODIC = European mouse disease clinics http://www.europhenome.org/

• CA – NorComm http://www.norcomm.org• J – RIKEN BRC Japan Mouse Clinic http://www.brc.riken.jp/lab/jmc/mouse_clinic/en/

RESOURCES: Find Phenotype Data and Protocols (Mice)

72

• For REPORTING (publishing) Animal Research: NIH.gov ‐ summary list (incomplete)

o https://www.nlm.nih.gov/services/research_report_guide.html

ARRIVE guidelines – Kilkenny & al 2010 – NC3R’s o https://www.ncbi.nlm.nih.gov/pubmed/20613859

ILAR/NAS guidance for reporting (NRC 2011)o http://www.nap.edu/catalog.php?record_id=13241

FASEB Recommendations 2016 o https://www.faseb.org/Portals/2/PDFs/opa/2016/FASEB_Enhancing%20Research%20Reproducibility.pdf

RESOURCES: (Guidelines)

73

• For PROTOCOLS and PLANNING studies Protocols.io: Virtual Communities for Protocol Development and Discussion. Teytelman et al. 2016.

o https://www.ncbi.nlm.nih.gov/pubmed/27547938o Find, discuss, implement, report, revise protocols, ‘publish’ with DOI https://www.protocols.io/

PREPARE: Guidelines for planning animal research and testing. Smith et al. 2017.

o https://www.ncbi.nlm.nih.gov/pubmed/28771074

RESOURCES: (Other)

74

Guidelines, Guidance, Recommendations: For REPORTING/REVIEWING:

ARRIVE Guidelines; Kilkenny 2010 https://www.nc3rs.org.uk/sites/default/files/documents/Guidelines/NC3Rs%20ARRIVE%20Guidelines%20Checklist%20(fillable).pdf

ILAR guidance NRC 2011http://ilarjournal.oxfordjournals.org/content/55/3/536.full.pdf+html

NIH Principles and Guidelines for Reporting Preclinical Research 2014 http://www.nih.gov/research‐training/rigor‐reproducibility/principles‐guidelines‐reporting‐preclinical‐research

FASEB Recommendations 2016 Enhancing Research Reproducibility https://www.faseb.org/Portals/2/PDFs/opa/2016/FASEB_Enhancing%20Research%20Reproducibility.pdf

etc

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Which mouse A or B is a better candidate recipient for a xenograft ?

A. Crl:CD1‐Foxn1nu

B.BALB/cAn ‐Foxn1nuNCr

Sex F FAge 6w 6mWBC 3x103/ul 6x103/ulTP 4 6Alb 3 3

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a) Why?

b) Which is out bred ?



immune variation disclosures resources and research impacts rml... · 2018-08-06 · presentation...

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