PDF《Supplementary Appendix》

Supplementary Appendix This appendix has been provided by the authors to give readers additional information about their work.

Supplement to: Lupski JR, Reid JG, Gonzaga-Jauregui C, et al. Whole-genome sequencing in a patient with CharcotCMarieCTooth neuropathy. N Engl J Med 2010;

362:1181-91. DOI: 10.1056/NEJMoa0908094. Supplementary?Appendix?to:? WholeGenome?Sequencing?and?SH3TC2?Mutations?Causing?CharcotMarieTooth?Neuropathy? James?R.?Lupski,?M.D.,?Ph.D.,?Jeffrey?G.?Reid,?Ph.D.,?Claudia?GonzagaJauregui,?B.S.,?David?Rio? Deiros,?B.S.,?David?C.Y.?Chen,?M.Sc.,?Lynne?Nazareth,?Ph.D.,?Matthew?Bainbridge,?M.Sc.,?Huyen? Dinh,?B.S.,?Chyn?Jing,?M.Sc.,?David?A.?Wheeler,?Ph.D.,?Amy?L.?McGuire,?J.D.,?Ph.D.,?Feng?Zhang,? Ph.D.,?Pawel?Stankiewicz,?M.D.,?Ph.D.,?John?J.?Halperin,?M.D.,?Chengyong?Yang,?Ph.D.,?Curtis? Gehman,?Ph.D.,?Danwei?Guo,?M.Sc.,?Rola?K.?Irikat,?B.S.,?Warren?Tom,?B.S.,?Nick?J.?Fantin,?B.S.,? Donna?M.?Muzny,?M.Sc.,?Richard?A.?Gibbs,?Ph.D.? From?the?Department?of?Molecular?and?Human?Genetics?(J.R.L.,?J.G.R.,?C.G.J.,?M.B.,?F.Z.,?P.S.,? D.M.M.,?R.A.G),?Human?Genome?Sequencing?Center?(J.G.R,?D.R.D.,?D.C.Y.C.,?L.N.,?M.B.,?H.D.,?C.? J.,?D.A.W.,?D.M.M.,?R.A.G),?Center?for?Medical?Ethics?and?Health?Policy?(A.L.M.),?Department?of? Pediatrics?(J.R.L.),?and?Texas?Children'

s?Hospital?(J.R.L.),?Baylor?College?of?Medicine,?Houston;

? Atlantic?Neuroscience?Institute,?Summit,?New?Jersey?and?Mount?Sinai?School?of?Medicine,?New? York?(J.J.H.);

?Life?Technologies,?Carlsbad?(C.Y.,?C.G.,?D.G.,?R.K.I.,?W.T.,?N.J.F)? Address?reprint?requests?to?Dr.?Richard?Gibbs?at?One?Baylor?Plaza,?Mail?Stop?BCM226,?Houston,? TX,?77030,[email protected].? ? Supplementary?Material?Contents:? 1. Supplementary?Methods,? 2. Supplementary?Figure?1.?Raw?read?coverage?at?key?SNP?positions.? 3. Supplementary?Table?1.?CNVs?found?in?neuropathy?subject'

s?genome? 4. Supplementary?Table?2.?nsSNPs?crossreferenced?to?the?Human?Gene?Mutation? Database? 5. Supplementary?Table?3.?Neuropathy?associated?genes.? ? 1. Supplementary?Methods? ? The?Applied?Biosystems?SOLiD??System?DNA?sequencing?platform?was?employed?to?sequence?genomic? DNA,?according?to?manufacturer'

s?specifications.??Primary?sequence?analysis?consisting?of?image?analysis? and?basecalling?was?performed?using?the?standard?SOLiD?primary?analysis?pipeline?for?the?35bp? fragment?and?25x25bp?matepair?data,?mapping?and?variant?calling?analysis?was?with?the?ABprovided? offline?analysis?suite? corona_lite .??For?the?50?x?50?matepair?data?the?Applied?Biosystems?Hidra? pipeline?was?employed.? Mapping?parameters?for?the?35?bp?reads?allowed?for?up?to?2?mismatches?in?25bp,?and?3?mismatches?in? 35bp.??The?mapping?did?not?allow?for?insertions?or?deletions.??Variant?calls?were?generated?in?a?high? sensitivity/low?stringency?mode?where?no?coverage?cutoffs?were?imposed?for?variant?calls.??Also,?multi allelic?sites?are?rejected?as?sequencing?noise,?or?mismapping.??In?making?SNP?calls,?multiple?reads?with? the?same?start?point?and?direction?are?treated?as?a?single?read?to?avoid?errors?from?oversampling?of? duplicate?read?alleles.?? To?maximize?the?sequence?yield?of?the?50x50bp?matepair?data?mapping?was?performed?in?28?different? iterations?with?reads?trimmed?back?to?progressively?shorter?lengths?(in?5bp?increments?from?50bp?down? to?25bp)?and?lower?stringency?(from?0?to?4?mismatches)( progressive?mapping ).??For?any?given?read?the? longest?mapping?with?the?highest?stringency?(longest?length,?fewest?mismatches)?was?kept?as?the?best? alignment.??Mapping?was?carried?out?with?mapreads?(the?mapping?tool?provided?by?the?vendor?as?part?of? their? corona_lite ?analysis?suite)?in?parallel?for?the?different?iterations?and?read?data?was?merged?at?the? end.? CNV?analysis?was?performed?with?the?AB?singlesample?CNV?tool?that?uses?read?coverage?corrected?for? mapping?and?amplification?efficiency?to?identify?regions?likely?to?have?increased?or?decreased?copy? number?variation.??Analysis?of?the?highdensity?random?array?data?used?custom?tools?developed?by? Applied?Biosystems?and?included?modified?beadfinding?and?progressive?mapping?using? corona_lite .?? Only?publicly?available?(www.solidsoftwaretools.com)?vendor?software?was?used?for?the?sequence?data? analysis?presented?in?this?study.? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? G

231123231302030230100112 A

311210313020302301001123 T

232313020302301001123103 T

0222233323202001231231310021133120312220032231123231302030230100112310313013002133011002202030131 TTCTCTATAGCTTCCCAGCAGCATGGGACATACTTACTCTTTAGATGTCGATGCCTTAAGCCAAACAGCAATGCCATTTCATAACAAAGAAGGCCATG AAGAGATATCGAAGGGTCGTCGTACCCTGTATGAATGAGAAATCTACAGCTACGGAATTCGGTTTGTCGTTACGGTAAAGTATTGTTTCTTCCGGTAC 0222233323202001231231310021133120312220032231123231302030230100112310313013002133011002202030131C 1331203122200322311232313020302301T 003223112323130203023010T A C

2223332320200123123131002113312031222003223112103 C

2020012312313100211331203222200322311200313020302 C

0112312313100211331203222200322311232313020122301 C

0123123131002113312031222003223112103130203023010 G 31310021113120312220032231121031302030230102..... C

1310021133120312220032231121031302030230100112310 C

133120312220032231121031 C

1331203122200322311210313020302301001123103130130 C 133120312220032231121031302030212100112. C

1331203122200322311210313020302301001123103130130 G

231123231302030230100112 G

231123231302030230100112 A

311210313020302301001123 A

311232313020302301001123 T

232313020302301001123103 G 2313020302301001123103130130021330110022020301… T

0222233323202001231231310021133120312220032231123231302030230100112310313013002133011002202030131 TTCTCTATAGCTTCCCAGCAGCATGGGACATACTTACTCTTTAGATGTCGATGCCTTAAGCCAAACAGCAATGCCATTTCATAACAAAGAAGGCCATG AAGAGATATCGAAGGGTCGTCGTACCCTGTATGAATGAGAAATCTACAGCTACGGAATTCGGTTTGTCGTTACGGTAAAGTATTGTTTCTTCCGGTAC 0222233323202001231231310021133120312220032231123231302030230100112310313013002133011002202030131C 1231231010021133100312220032231121T 2222333232020012312313100212331203122200322311232T 2320200123123131002113312031222003223112323130203T 2020012312313100211331203122200322311210313020302C 1231231310021133120312220032231123231302030230100T 1310021133120312220032231121031302030230100112310T 31002113312........