Size-reduction of a broad host-range BioBrick cloning vector
Andreas Constantinou
Degree project in biology, Bachelor of science, 2010 Examensarbete i biologi 15 hp till kandidatexamen, 2010
Biology Education Centre, Uppsala University and Department of Photochemistry and Molecular Science
Supervisors: Thorsten Heidorn and Daniel Camsund
6XPPDU\
7KH %LR%ULFN DVVHPEO\ VWDQGDUG LV D FORQLQJ VWDQGDUG GHYHORSHG IRU IDFLOLWDWLQJ WKH DVVHPEO\ DQG FORQLQJ RI VWDQGDUGL]HG JHQHWLF SDUWV %LR%ULFNV IRU EXLOGLQJ QHZ ELRORJLFDO FLUFXLWV 7R HQDEOH %LR
%ULFN H[SUHVVLRQ LQ F\DQREDFWHULD D QHZ PRELOL]DEOH %LR%ULFN YHFWRU EDVHG RQ WKH EURDGKRVWUDQJH SODVPLG 56) ZDV UHFHQWO\ GHYHORSHG DQG QDPHG S304$. 7R LPSURYH WKH FORQLQJ FDSDF
LW\ RI WKH YHFWRU , KDYH VWXGLHG WKH 56) UHSOLFRQ DQG LGHQWLILHG JHQHWLF HOHPHQWV WKDW DUH HLWKHU GLVSRVDEOH RU H[FKDQJHDEOH 6HYHUDO %LR%ULFNV ZHUH GHVLJQHG DQG FUHDWHG IURP WKH 56) UHSOL
FRQ DQG IURP WKH 53 SODVPLG WR SUHSDUH WKH FRQVWUXFWLRQ RI D VL]HUHGXFHG PRELOL]DEOH YHUVLRQ RI S304$. )XUWKHUPRUH IRXU %LR%ULFN DQWLELRWLF UHVLVWDQFH FDVVHWWHV ZHUH H[WUDFWHG DQG WHVWHG IRU H[SUHVVLRQ RI UHVLVWDQFH WR DPSLFLOOLQ NDQDP\FLQ FKORUDPSKHQLFRO DQG WHWUDF\FOLQH LQ (VFKHULFKLD FROL D %LR%ULFN HQFRGLQJ UHVLVWDQFH WR VSHFWLQRP\FLQ ZDV DOVR FUHDWHG 7KHVH PD\ ODWHU EH XVHG DV VHOHFWLYH PDUNHUV LQ S304$. DQG RWKHU %LR%ULFN YHFWRUV
,QWURGXFWLRQ
%LR%ULFN VWDQGDUG DVVHPEO\
,Q WKH SDVW GHFDGH V\QWKHWLF ELRORJ\ KDV HPHUJHG DV D QHZ SURPLVLQJ ILHOG RI UHVHDUFK DLPLQJ WRZDUG XVLQJ WKH FRPELQHG NQRZOHGJH LQ PROHFXODU ELRORJ\ WR JHQHWLFDOO\ HQJLQHHU QHZ ELRORJLFDO SDUWV
GHYLFHV DQG V\VWHPV DV ZHOO DV WR LPSURYH DQG PRGLI\ H[LVWLQJ ELRORJLFDO V\VWHPV IRU XVHIXO SXUSRVHV KWWSV\QWKHWLFELRORJ\RUJ
7R IDFLOLWDWH WKH FRPELQDWLRQ RI JHQHWLF SDUWV LQWR FRPSOH[ JHQHWLF FLUFXLWV D '1$ DVVHPEO\ VWDQ
GDUG ZDV SURSRVHG E\ 7RP .QLJKW DW WKH 0DVVDFKXVHWWV ,QVWLWXWH RI 7HFKQRORJ\ LQ .QLJKW
'XEEHG %LR%ULFN VWDQGDUG DVVHPEO\ WKLV VWDQGDUG GHVFULEHV D VWUHDPOLQHG SURFHVV IRU DVVHP
EO\ DQG FORQLQJ RI DQ\ WZR RU PRUH VWDQGDUGL]HG JHQHWLF SDUWV LQWR VWDQGDUGL]HG SODVPLG YHFWRUV
(DFK VWDQGDUGL]HG SDUW RU %LR%ULFN LV IODQNHG E\ D ¶ VWDQGDUG SUHIL[ VHTXHQFH ¶ JDDWWFJFJJF
FJFWWFWDJ ¶ IRU SDUWV VWDUWLQJ ZLWK DWJ ¶ JDDWWFJFJJFFJFWWFWDJDJ ¶ IRU DOO RWKHU SDUWV FRQWDLQLQJ (FR5, DQG ;ED, UHVWULFWLRQ VLWHV DQG D ¶ VWDQGDUG VXIIL[ VHTXHQFH ¶ WDFWDJWDJFJJFFJFWJFDJ ¶
FRQWDLQLQJ 6SH, DQG 3VW, UHVWULFWLRQ VLWHV 6LPLODUO\ VWDQGDUGL]HG %LR%ULFN SODVPLGV GHVLJQHG ZLWK LGHQWLFDO SUHIL[ DQG VXIIL[ VHTXHQFHV DUH XVHG DV WDUJHW FORQLQJ YHFWRUV )RU DVVHPEO\ RI D VLQJOH
%LR%ULFN SDUW LQWR D %LR%ULFN SODVPLG YHFWRU ERWK SDUW DQG YHFWRU DUH GLJHVWHG ZLWK (FR5, DQG 3VW, UHVWULFWLRQ HQ]\PHV WKLV FUHDWHV FRKHVLYH HQGV WKDW PD\ EH OLJDWHG WR IRUP D FORVHG FLUFXODU SODVPLG QRZ FRQWDLQLQJ WKH %LR%ULFN SDUW DV DQ LQVHUW +RZHYHU WKH UHDO VWUHQJWK RI %LR%ULFN DVVHPEO\ LV UH
YHDOHG ZKHQ FRPELQLQJ WZR RU PRUH %LR%ULFN SDUWV WRJHWKHU $VVHPEO\ RI WZR %LR%ULFNV LV SUHSDUHG E\ GLJHVWLQJ RQH SDUW ZLWK 6SH, DQG WKH RWKHU ZLWK ;ED, 'LJHVWHG 6SH, DQG ;ED, VLWHV IRUP FRKHVLYH HQGV ZKHQ OLJDWHG WRJHWKHU WKH\ IRUP D VFDU VHTXHQFH ZKLFK LV LQGLJHVWLEOH E\ HLWKHU 6SH, RU ;ED,
$V D UHVXOW WKH WZR DVVHPEOHG %LR%ULFNV FDQ QRZ EH UHJDUGHG DV D FRPSRVLWH SDUW DQG PD\ EH HLWKHU DVVHPEOHG LQWR D %LR%ULFN SODVPLG YHFWRU RU IXUWKHU FRPELQHG ZLWK DGGLWLRQDO %LR%ULFNV ,Q SUDFWLFH
DVVHPEO\ RI WZR SDUWV LQWR D YHFWRU PD\ EH GRQH LQ SDUDOOHO )LJXUH .QLJKW 6KHWW\ HW DO
%LR%ULFN SDUWV DQG YHFWRUV DUH PDGH UHDGLO\ DYDLODEOH LQ D GHGLFDWHG '1$ UHJLVWU\ FDOOHG WKH 5HJLVWU\ RI 6WDQGDUG %LRORJLFDO 3DUWV 5HJLVWU\ KWWSSDUWVUHJLVWU\RUJ
%URDGKRVWUDQJH %LR%ULFN SODVPLG S304$.
:LWK WKHLU DELOLW\ WR FRQYHUW OLJKW WR FKHPLFDO HQHUJ\ F\DQREDFWHULD KDYH UHFHLYHG PXFK DWWHQWLRQ LQ WKH ILHOG RI V\QWKHWLF ELRORJ\ DV IXWXUH ELRORJLFDO IDFWRULHV IRU SURGXFWLRQ RI QH[WJHQHUDWLRQ ELRIXHOV 'XWWD HW DO +XDQJ HW DO KWWSZZZV\QWKHWLFJHQRPLFVFRP +RZHYHU PRVW JHQHWLF WRROV XVHG LQ WKH %LR%ULFN DVVHPEO\ VWDQGDUG KDYH EHHQ GHYHORSHG IRU FORQLQJ LQ (VFKHULFKLD FROL
7KHUHIRUH WR DOORZ WKH XVH RI WKH %LR%ULFN DSSURDFK LQ F\DQREDFWHULD +XDQJ HW DO UHFHQWO\
GHYHORSHG D QHZ EURDGKRVWUDQJH %LR%ULFN YHFWRU 7KH YHFWRU ZDV FRQVWUXFWHG E\ IXVLQJ WKH UHSOL
FRQ IURP WKH EURDGKRVWUDQJH SODVPLG 56) ZLWK WKH %LR%ULFN SODVPLG S6%$. IURP ZKLFK WKH UHSOLFRQ KDG EHHQ UHPRYHG 7KLV UHVXOWHG LQ D QHZ EDVH SDLU ES %LR%ULFN SODVPLG YHF
WRU S304$. FDSDEOH RI UHSOLFDWLQJ LQ ERWK ( FROL DQG F\DQREDFWHULD )LJXUH )XUWKHUPRUH
WKH SODVPLG LV DOVR PRELOL]DEOH WKURXJK FRQMXJDWLRQ ZLWK WKH KHOS RI D KHOSHU SODVPLG VXFK DV 53
DOORZLQJ LW WR EH WUDQVIHUUHG WR QRQWUDQVIRUPDEOH FHOOV +XDQJ HW DO
'HVSLWH WKHVH DGYDQWDJHV RQH GUDZEDFN RI S304$. LV LWV VL]H ZKLFK PD\ EH LOOXVWUDWHG E\
FRPSDULQJ LW WR WKH ES %LR%ULFN YHFWRU S6%$. IURP ZKLFK LW ZDV GHULYHG $V WKH VL]H RI D SODVPLG LQFUHDVHV VR GRHV LWV PHWDEROLF LPSDFW RQ LWV KRVW ,I WKLV LPSDFW JHWV WRR ODUJH WKH SODVPLG PLJKW EHFRPH XQVWDEOH LH GLIILFXOW IRU WKH FHOO WR PDLQWDLQ FDXVLQJ WKH FHOO WR ORVH WKH SODVPLG ,Q FORQLQJ LW LV WKHUHIRUH GHVLUDEOH WR NHHS WKH VL]H RI D YHFWRU WR D PLQLPXP WR PD[LPL]H WKH VWDELOLW\
DQG FDSDFLW\ RI WKH SODVPLG HYHQ DV QHZ JHQHWLF PDWHULDO LV DGGHG
)LJXUH $VVHPEO\DQGOLJDWLRQRIWZR%LR%ULFNSDUWVLQWRD%LR%ULFNYHFWRU 7KHWZRSDUWV GHQRWHGSUHÀ[DQGVXIÀ[
DUHHDFKFDUULHGRQVHSDUDWH%LR%ULFNYHFWRUV$ DQG%7KHSUHÀ[SDUWLVH[FLVHGE\GLJHVWLRQZLWK(FR5, DQG6SH,
ZKLOHWKHVXIÀ[SDUWLVH[FLVHGZLWK;ED, DQG3VW,6SH, DQG;ED, GLJHVWLRQUHVXOWVLQFRKHVLYHHQGVWKDWFDQEHOLJDWHG WRJHWKHUWRIRUPDVFDUVHTXHQFH LQGLJHVWLEOHE\HLWKHURIWKHWZRHQ]\PHV 0HDQZKLOH WKH%LR%ULFNGHVWLQDWLRQYHFWRU & LVGLJHVWHGZLWK(FR5, DQG3VW,H[FLVLQJWKHFFG% LQVHUWFRGLQJIRUD'1$ J\UDVHWR[LQ 'LJHVWHGGHVWLQDWLRQYHFWRU LVOLJDWHGZLWKWKHFRPSRVLWHSUHÀ[VXIÀ[SDUW'$IWHUWUDQVIRUPDWLRQ WKHGHVWLQDWLRQYHFWRULVSRVLWLYHO\VHOHFWHG IRUE\JURZLQJFHOOVZLWKWKHDQWLELRWLFFRUUHVSRQGLQJWRWKHYHFWRUҋVXQLTXHDQWLELRWLFUHVLVWDQFHPDUNHU 9HFWRUVUH
OLJDWHGZLWKFFG% LQVHUWVDUHDXWRPDWLFDOO\VHOHFWHGDJDLQVWE\XVLQJDWR[LQVHQVLWLYHVWUDLQ 6LQFH(FR5, DQG3VW, GRQҋW SURGXFHFRPSOHPHQWDU\RYHUKDQJV YHFWRUVQRWFDUU\LQJDQ\LQVHUWZLOOUHPDLQOLQHDU $EEUHYLDWLRQV ((FR5,66SH,
;;ED,33VW,$PS5$PSLFLOOLQUHVLVWDQFHFDVVHWWH &P5&KORUDPSKHQLFROUHVLVWDQFHFDVVHWWH.P5.DQDP\FLQ UHVLVWDQFHFDVVHWWH )LJXUHDGDSWHGDQGUHSURGXFHGIURP6KHWW\ HW DO XQGHUWKH&UHDWLYH&RPPRQV$WWULEXWLRQ /LFHQVHKWWSFUHDWLYHFRPPRQVRUJOLFHQVHVE\
7KH 56) SODVPLG UHSOLFRQ
7KH 56) SODVPLG ZDV RULJLQDOO\ LVRODWHG IURP 6DOPRQHOOD HQWHULFD LQ 0H\HU DQG KDV VLQFH EHHQ VKRZQ WR UHSOLFDWH VXFFHVVIXOO\ LQ D ZLGH UDQJH RI ERWK JUDPQHJDWLYH DQG JUDPSRVLWLYH EDFWHULD LQFOXGLQJ ( FROL DQG PHPEHUV RI WKH F\DQREDFWHULDO JHQHUD 6\QHFKRFRFFXV 6\QHFKRF\VWLV DQG 1RVWRF +XDQJ HW DO 0HUPHW%RXYLHU HW DO 0H\HU 6RGH HW DO $O
WKRXJK QRQVHOIWUDQVPLVVLEOH LH XQDEOH WR DORQH WULJJHU FRQMXJDO WUDQVIHU IURP RQH FHOO WR DQRWKHU
56) KDV HYROYHG DQ DGYDQFHG V\VWHP IRU SDUDVLWL]LQJ RQ WKH FRQMXJDWLRQ V\VWHPV RI PDQ\ RWKHU SODVPLG W\SHV VXFK DV WKH FRQMXJDWLYH SODVPLG 53 0H\HU
56) UHSOLFDWLRQ
$W OHDVW WKUHH JHQHV KDYH EHHQ LGHQWLILHG DV QHFHVVDU\ IRU UHSOLFDWLRQ RI WKH 56) SODVPLG 7KHVH DUH WKH UHSOLFDWLRQ JHQHV UHS$ UHS% DQG UHS& HQFRGLQJ D '1$ KHOLFDVH D SULPDVH DQG D UHSOLFDWLRQ LQLWLDWLRQ SURWHLQ UHVSHFWLYHO\ 7KH JHQHV DUH FOXVWHUHG WRJHWKHU DQG FRWUDQVFULEHG VHSDUDWHG RQO\ E\
WZR VPDOO JHQHV HQFRGLQJ D VPDOO UHSUHVVRU SURWHLQ 5HS) DQG D SURWHLQ ZLWK DQ XQNQRZQ IXQFWLRQ 5HS( )LJXUH ,QWHUHVWLQJO\ WKH SULPDVH LV H[SUHVVHG LQ WZR GLIIHUHQW IRUPV DORQH GHQRWHG 5HS%¶ DQG LQ IXVLRQ ZLWK WKH 0RE$ SURWHLQ VHH EHORZ WKHQ UHIHUUHG WR DV 5HS%
5HS$ 5HS% DQG 5HS& LQLWLDWH SODVPLG UHSOLFDWLRQ E\ IRUPLQJ D UHSOLVRPH DW WKH YHJHWDWLYH UHSOL
FDWLRQ RULJLQ RUL9 7KH ILUVW VWHS LQ WKLV SURFHVV LQYROYHV WKH UHFRJQLWLRQ RI LQYHUWHG UHSHDWV RU
)LJXUH 7KH%LR%ULFNEURDGKRVWUDQJHSODVPLGYHFWRUS304$. 7KHSODVPLGZDVFRQVWUXFWHGE\FRPELQLQJWKH EURDGKRVWUDQJH56)SODVPLGUHSOLFRQZLWKWKH%LR%ULFNSODVPLGS6%$. IURPZKLFKWKHRULJLQDOUHSOLFRQKDG EHHQUHPRYHG ,QFRQWUDVWWRRWKHU%LR%ULFNYHFWRUVDYDLODEOHDWWKHWLPH S304$.LVVXFFHVVIXOO\UHSOLFDWHGLQERWK (FROL DQGVHYHUDOF\DQREDFWHULDOVWUDLQV DVZHOODVPRELOL]DEOHWKURXJKFRQMXJDWLRQ $EEUHYLDWLRQVGHVFULEHGLQERG\
WH[W )LJXUHJHQHUDWHGZLWK*HQHLRXV3URELRLQIRUPDWLFVVRIWZDUHKWWSZZZJHQHLRXVFRP
LWHURQV LQ WKH RUL9 UHJLRQ E\ 5HS& 8SRQ ELQGLQJ 5HS& GLVUXSWV WKH KHOLFLW\ RI WKH SODVPLG '1$
DOORZLQJ HQWU\ RI WKH 5HS$ KHOLFDVH ZKLFK LQ WXUQ VHSDUDWHV WKH WZR '1$ VWUDQGV IRU HQWU\ RI 5HS%
5HS% WKHQ SULPHV SODVPLG UHSOLFDWLRQ SHUPLWWLQJ V\QWKHVLV RI '1$ E\ WKH KRVWFHOO '1$ SRO\PHUDVH ,,, 0H\HU
56) PRELOL]DWLRQ
56) LV PRELOL]HG E\ SURWHLQV HQFRGHG E\ WKUHH JHQHV FDOOHG PRELOL]DWLRQ PRE JHQHV $ % DQG &
7KHVH DUH ORFDWHG LQ D UHJLRQ VXUURXQGLQJ WKH SODVPLG¶V RULJLQ RI WUDQVIHU RUL7 IURP ZKLFK SODVPLG WUDQVIHU LV LQLWLDWHG )LJXUH $V DOUHDG\ GHVFULEHG PRE$ LV WUDQVODWHG WRJHWKHU ZLWK UHS% WR IRUP WKH IXVLRQ SURWHLQ 5HS% LQ ZKLFK 0RE$ PDNHV DQ 1WHUPLQDO GRPDLQ $OVR RYHUODSSLQJ WKH PRE$
DQG UHS% UHJLRQ LV WKH PRE% JHQH KRZHYHU 0RE% LV WUDQVODWHG LQ D GLIIHUHQW UHDGLQJ IUDPH WKDQ 5HS% 7KH WKLUG PRE JHQH PRE& LV ORFDWHG RQ WKH RSSRVLWH VLGH RI RUL7 DQG LV WKH RQO\ JHQH RI WKH 56) UHSOLFRQ WUDQVFULEHG DQG WUDQVODWHG LQ WKH RSSRVLWH GLUHFWLRQ
6LPLODU WR WKH UHSOLFDWLRQ SURWHLQV 0RE$ 0RE% DQG 0RE& FRPH WRJHWKHU WR IRUP D IXQFWLRQDO XQLW FDOOHG WKH UHOD[RVRPH 7KH SXUSRVH RI WKH UHOD[RVRPH LV WR LQLWLDWH WKH RSHQLQJ DQG WUDQVIHU RI WKH SODVPLG DV ZHOO DV WR WDUJHW LW WR DQ DYDLODEOH FRQMXJDWLRQ V\VWHP 7KH UHOD[RVRPH DOVR DFWV DV D WUDQVFULSWLRQDO UHSUHVVRU DV LV GHVFULEHG EHORZ 0H\HU
7UDQVFULSWLRQDO UHJXODWLRQ RI 56) JHQHV
0RVW RI WKH WUDQVFULSWLRQ LQ WKH 56) UHSOLFRQ VWDUWV IURP WKUHH SURPRWHUV ORFDWHG DGMDFHQW WR RUL7 )LJXUH 7KHVH SURPRWHUV GHQRWHG S S DQG S DUH VR QDUURZO\ VSDFHG WKDW WKHLU DQG
UHJLRQV RYHUODS $V D UHVXOW WKH\ DUH DOO VLPXOWDQHRXVO\ UHJXODWHG E\ WKH VDPH UHSUHVVRU FRPSOH[
S DQG S WUDQVFULEH FORFNZLVH DV VKRZQ LQ ILJXUH WKURXJK RUL7 UHS% DQG WKH UHS$& RSHURQ ,Q FRQWUDVW WKH S SURPRWHU ORFDWHG EHWZHHQ S DQG S WUDQVFULEHV FRXQWHUFORFNZLVH DQG LV WKH VROH SURPRWHU WUDQVFULELQJ PRE&
)LJXUH 2UJDQL]DWLRQDQGUHJXODWLRQRIJHQHH[SUHVVLRQ UHSOLFDWLRQDQGWUDQVIHULQWKH56)UHSOLFRQ )LOOHG DUURZVDQGER[HVUHSUHVHQWSURWHLQFRGLQJJHQHVDQGRWKHUJHQHWLFHOHPHQWV UHVSHFWLYHO\ 'DVKHGDUURZVUHSUHVHQW WUDQVFULSWLRQGLUHFWLRQIURPSURPRWHUV $EEUHYLDWLRQVGHVFULEHGLQERG\WH[W )LJXUHEDVHGRQSUHYLRXVZRUNE\0H\HU
'XH WR WKHLU ORFDWLRQ LQ FORVH YLFLQLW\ RI RUL7 SURPRWHUV SS DUH LQGLUHFWO\ DQG QHJDWLYHO\ UH
SUHVVHG E\ WKH ELQGLQJ RI WKH UHOD[RVRPH WR RUL7 &RQVHTXHQWO\ H[SUHVVLRQ RI 0RE$ 5HS% DQG 0RE% IURP WKH S DQG S SURPRWHUV UHVXOWV LQ D FRPSOH[ VHOIUHJXODWLQJ QHJDWLYH IHHGEDFN ORRS
,Q DGGLWLRQ WR WKHVH D IRXUWK SURPRWHU GHQRWHG S LV ORFDWHG GLUHFWO\ GRZQVWUHDP RI UHS% S
WUDQVFULEHV WKH UHS$& RSHURQ WKURXJK WKH SUHYLRXVO\ GHVFULEHG JHQHV UHS( DQG UHS) )LJXUH 7KH ODWWHU RI WKHVH HQFRGHV D VPDOO UHSUHVVRU SURWHLQ ZKLFK QHJDWLYHO\ UHJXODWHV S S DQG UHS) FDQ EH GHOHWHG ZLWKRXW DIIHFWLQJ WKH SODVPLG FRS\QXPEHU +RZHYHU LW LV EHOLHYHG WKDW WKH S FRQWURO V\VWHP KHOSV LQFUHDVH WKH SODVPLG¶V UDWH RI HVWDEOLVKPHQW XSRQ DUULYDO LQ D QHZ KRVW DV LW DOORZV IRU HDUO\
H[SUHVVLRQ RI WKH UHS$ DQG UHS% JHQHV 0H\HU
%LR%ULFN DQWLELRWLF UHVLVWDQFH FDVVHWWHV
0RVW SODVPLG YHFWRUV XVHG IRU FORQLQJ DUH HTXLSSHG ZLWK VHOHFWLYH PDUNHUV WR DOORZ IRU VHOHFWLRQ RI FHOOV FDUU\LQJ WKH YHFWRU WKHVH PDUNHUV DUH RIWHQ JHQHV H[SUHVVLQJ UHVLVWDQFH WR YDULRXV DQWLELRWLFV
7KH S304$. %LR%ULFN YHFWRU FDUULHV WZR VXFK DQWLELRWLF UHVLVWDQFH FDVVHWWHV H[SUHVVLQJ UHVLVWDQFH WR DPSLFLOOLQ $PS5 DQG NDQDP\FLQ .P5 +XDQJ HW DO 'LIIHUHQW VLWXDWLRQV PD\ KRZHYHU
UHTXLUH GLIIHUHQW FRPELQDWLRQV RI VHOHFWLYH PDUNHUV ,W LV WKHUHIRUH RIWHQ GHVLUDEOH WR KDYH YDULRXV YHFWRUV HDFK H[SUHVVLQJ D GLIIHUHQW VHW RI DQWLELRWLF UHVLVWDQFHV 7KH %LR%ULFN DVVHPEO\ VWDQGDUG DOORZV DOO %LR%ULFN YHFWRUV WR EH PRGLILHG DQG HTXLSSHG ZLWK DQ\ FRPELQDWLRQ RI VHOHFWLYH PDUNHUV DYDLODEOH DV %LR%ULFNV 6KHWW\ HW DO
$LPV
,Q WKLV VWXG\ , LQYHVWLJDWHG WKH 56) UHSOLFRQ IRU JHQHV DQG UHJLRQV WKDW DUH GLVSHQVDEOH IURP WKH S304$. %LR%ULFN FORQLQJ YHFWRU ZLWKRXW ORVLQJ LWV NH\ IHDWXUHV DV D EURDG KRVWUDQJH DQG PRELOL]DEOH SODVPLG , DOVR DWWHPSWHG D VWDQGDUGL]DWLRQ RI WKH HQWLUH 56) UHSOLFRQ DFFRUGLQJ WR
WKH %LR%ULFN VWDQGDUG
)XUWKHUPRUH WR HQDEOH PRGLILFDWLRQ RI WKH DQWLELRWLF UHVLVWDQFH UHSHUWRLUH RI S304$. IRXU DQ
WLELRWLF UHVLVWDQFH FDVVHWWH %LR%ULFNV REWDLQHG IURP WKH 5HJLVWU\ FDWDORJXH ZHUH WHVWHG IRU H[SUHVVLRQ RI DQWLELRWLF UHVLVWDQFH DQG D QHZ %LR%ULFN HQFRGLQJ UHVLVWDQFH WR D ILIWK DGGLWLRQDO DQWLELRWLF ZDV FRQVWUXFWHG
5HVXOWV
%LR%ULFN VWDQGDUGL]DWLRQ DQG VL]HUHGXFWLRQ RI WKH 56) UHSOLFRQ
'HVLJQ RI D VL]HUHGXFHG 56) UHSOLFRQ
,W KDV EHHQ VKRZQ WKDW GHOHWLRQ RI WKH HQWLUH 0RE UHJLRQ LQFOXGLQJ RUL7 DQG WKH SS SURPRWHUV
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
HVVHQWLDO . OQ WKH RWKHU KDQG WKH UHTXLUHPHQW fRr DQ 53 KHOSHU SODVPLG IRU 56) PRELOL]DWLRQ UDLVHG WKH TXHVWLRQ ZKHWKHU WKH WUDQVIHULQLWLDWLQJ UROH RI WKH 56) UHOD[RVRPH FDQ EH SURYLGHG LQ WUDQV IURP WKH KHOSHU SODVPLG
%DVHG RQ WKLV , K\SRWKHVL]HG WKH FRQVWUXFWLRQ RI D UHGXFHGVL]H YHUVLRQ WKH 56) UHSOLFRQ E\ UHPRYLQJ WKH HQWLUH 0RE UHJLRQ ≈ NE DQG UHSODFLQJ LW ZLWK WKH 53 RUL7 ≈ ES DV ZHOO DV ZLWK D QHZ SURPRWHU GULYLQJ H[SUHVVLRQ RI WKH UHS JHQHV )LJXUH ,Q WKHRU\ WKLV ZRXOG UHVXOW LQ D PRELOL]DEOH SODVPLG HQWLUHO\ GHSHQGHQW RQ 53 IRU FRQMXJDO WUDQVIHU LQGHHG LW KDV EHHQ VKRZQ WKDW WKH RUL7 LV WKH RQO\ 53 WUDQVIHU HOHPHQW UHTXLUHG LQ FLV 3DQVHJUDX HW DO 7KH UHVXOWLQJ SODVPLG YHFWRU ZRXOG QRW RQO\ EHQHILW IURP D UHGXFWLRQ LQ VL]H by XS WR ≈12 ES EXW ZRXOG DOVR KDYH DQ LQFUHDVHG YHFWRU VDIHW\ DV LWV SURSRVHG 53 GHSHQGHQFH ZRXOG PLQLPL]H WKH ULVN RI XQFRQWUROOHG SODVPLG VSUHDG
)LJXUH &RQVWUXFWLRQRIDUHGXFHGVL]HYHUVLRQRIS304$.ODFNLQJWKHPRELOL]DWLRQUHJLRQ 7KHVKDGHGÀHOGLQGL
FDWHVWKHPRELOL]DWLRQUHJLRQ≈ NESURSRVHGIRUUHSODFHPHQWE\ RUL7 IURPWKHFRQMXJDWLYHSODVPLG53≈ NE
$OVRPDUNHGDUHWKHUHJLRQVFRYHUHGE\WKHGHVLJQHG56)GHULYHG%LR%ULFNV $EEUHYLDWLRQVGHVFULEHGLQERG\
WH[W )LJXUHJHQHUDWHGZLWK*HQHLRXV3URELRLQIRUPDWLFVVRIWZDUHKWWSZZZJHQHLRXVFRP
)LJXUH *HOHOHFWURSKRUHVLVDQDO\VHVRIFRQVWUXFWHG%LR%ULFNVIRUUHGXFWLRQRIWKH56)UHSOLFRQ %LR%ULFNSDUWV FRQVWUXFWHGE\3&5 DPSOLÀFDWLRQIURPSODVPLGVS304$.%%,QF4B; RUS5/%%53BRUL7 ZHUHDQDO\]HG E\DJDURVHJHOHOHFWURSKRUHVLV 3DUWVL]HVLQFOXGHVHTXHQFHVLQWURGXFHGE\ҋSULPHUH[WHQVLRQVDQGDUHWKHUHIRUH ORQJHUWKDQWKHDFWXDO%LR%ULFNSDUWV D DJDURVH 9 PLQ OSHUVDPSOH Oλ E DJDURVH
9 PLQ OSHUVDPSOH Oλ λES 2ҋ*HQH5XOHU ES'1$ ODGGHUλNE 2ҋ*HQH5XOHU NE'1$
ODGGHU
'HVLJQ DQG FRQVWUXFWLRQ RI 56) DQG 53GHULYHG %LR%ULFN SDUWV
7R SUHSDUH WKH FRQVWUXFWLRQ DQG DVVHPEO\ RI D VL]HUHGXFHG YHUVLRQ RI S304$. VL[ %LR%ULFN SDUWV ZHUH GHVLJQHG 7KHVH LQFOXGHG ILYH SDUWV GHULYHG IURP WKH 56) UHSOLFRQ UHVSHFWLYHO\ FDUU\LQJ WKH S SURPRWHU %%,QF4B3 ES RUL9 %%,QF4BRUL9 ES WKH UHSJHQH UHJLRQ %%,QF4BUHS
ES RUL7 %%,QF4BRUL7 ES DQG WKH SS SURPRWHUV %%,QF4B33 ES )LJXUH
7KH VL[WK %LR%ULFN ZDV GHULYHG IURP WKH 53 SODVPLG FDUU\LQJ WKH 53 RUL7 %%53BRUL7
ES )RU %LR%ULFN VWDQGDUGL]DWLRQ RI WKH FXUUHQW FRPSOHWH 56) UHSOLFRQ RQH DGGLWLRQDO
%LR%ULFN FDUU\LQJ WKH HQWLUH 56) UHSOLFRQ %%,QF4B56) ES ZDV GHVLJQHG
3DUWV ZHUH FRQVWUXFWHG E\ SRO\PHUDVH FKDLQ UHDFWLRQ 3&5 '1$ DPSOLILFDWLRQ ZLWK IRUZDUG DQG UHYHUVH SULPHUV WKDW UHVSHFWLYHO\ LQWURGXFHG WKH %LR%ULFN SUHIL[ DQG VXIIL[ VHTXHQFHV DV ¶ SULPHU H[WHQVLRQV 7DEOH 56) SDUWV ZHUH GHVLJQHG DQG DPSOLILHG IURP WKH S304$. SODVPLG VH
TXHQFH *HQ%DQN DFF QR *8 7KH 53 SDUW ZDV GHVLJQHG IURP WKH '1$ VHTXHQFH RI SODVPLG ,QF3α *HQ%DQN DFF QR 1&B DQG DPSOLILHG IURP WKH 53GHULYDWLYH SODVPLG S5/ (OKDL DQG :RON 3&5 SURGXFWV ZHUH DQDO\]HG E\ DJDURVH JHO HOHFWURSKRUHVLV )LJXUH
ZKLFK UHYHDOHG VXFFHVVIXO DPSOLILFDWLRQ RI DOO GHVLJQHG SDUWV H[FHSW %%,QF4B56) DQG %%
,QF4B3
,Q DQ DWWHPSW WR WURXEOHVKRRW WKH XQVXFFHVVIXO 3&5 DPSOLILFDWLRQV RI %%,QF4B56) DQG %%
,QF4B3 DV ZHOO DV WR LGHQWLI\ WKH VRXUFH RI WKH ZHDN XQVSHFLILF EDQG REVHUYHG IRU %%,QF4BRUL9
UHOHYDQW SULPHUV ZHUH DOLJQHG DJDLQVW WKH QXFOHRWLGH VHTXHQFH RI S304$. XVLQJ WKH %DVLF /RFDO
$OLJQPHQW 6HDUFK 7RRO %/$67 KWWSEODVWQFELQOPQLKJRY WR LGHQWLI\ XQVSHFLILF SULPHU ELQGLQJ VLWHV 5HVXOWV UHYHDOHG WZR VXFK VLWHV IRU SULPHU )BRUL9%%SUH RI ZKLFK RQH ZDV LGHQWLFDO WR WKH LQWHQGHG DQQHDOLQJ VLWH PRUH VSHFLILFDOO\ WKH SULPHU ELQGV WR D UHJLRQ RI LQYHUWHG UHSHDWV $V D UHVXOW DW OHDVW WZR SURGXFWV GLIIHULQJ E\ RQO\ ES LQ VL]H FRXOG EH H[SHFWHG IURP WKH DPSOLILFDWLRQ RI ERWK %%,QF4B56) DQG %%,QF4BRUL9 )RU WKH ODWWHU WKLV ZRXOG \LHOG WKH GHVLUHG IXOOOHQJWK
ESSURGXFW DQG D WUXQFDWHG YHUVLRQ RI ES 'HVSLWH WKLV LW ZDV GHFLGHG WKDW QR UHGHVLJQ RI
)LJXUH &RORQ\3&5 YHULÀFDWLRQRIFORQHG56)DQG53%LR%ULFNV )RXUFORQHV$' RIHDFKVXFFHVVIXOO\FRQ
VWUXFWHGDQGFORQHG%LR%ULFN FDUULHGRQS6%$ ZHUHYHULÀHGE\FRORQ\3&5 IURP9)DQG95 SULPHUDQQHDOLQJVLWHV
ÁDQNLQJWKH%LR%ULFNLQVHUWVZLWK ES 3&5 SURGXFWVZHUHYHULÀHGIRUFRUUHFWVL]HVE\DJDURVHJHOHOHFWURSKRUH
VLV RQ DJDURVH JHOV DW 9 PLQ D DQG EDQG D DJDURVH JHO DW 9 PLQ %%,QF4BUHS
GDWDQRWVKRZQ ([SHFWHGIUDJPHQWVL]HV %%,QF4BRUL7 ES %%,QF4B33 ES %%53BRUL7 ES
%%,QF4BRUL9 ES %%,QF4BUHS ESλ ES 2ҋ*HQH5XOHU ES'1$ ODGGHU
)BRUL9%%SUH ZDV QHFHVVDU\ DV GHVLUHG IUDJPHQWV FRXOG EH LGHQWLILHG DW D ODWHU VWDJH E\ VHTXHQFLQJ FORQHG SDUWV 6HYHUDO DWWHPSWV ZHUH PDGH WR UHDPSOLI\ %%,QF4B56) DQG %%,QF4B3 XVLQJ YDULRXV 3&5 VHWWLQJV EXW DOO VXFK DWWHPSWV IDLOHG
6XFFHVVIXOO\ DPSOLILHG SDUWV ZHUH FORQHG LQWR SODVPLG YHFWRU S6%$ EHWZHHQ UHVWULFWLRQ VLWHV (FR5, DQG 3VW, DQG WKH UHVXOWLQJ SODVPLGV WUDQVIRUPHG LQWR ( FROL )RXU FORQHV $' RI HDFK FRQ
VWUXFW ZHUH WKHQ VHOHFWHG IRU DQDO\VLV RI LQVHUW OHQJWK E\ FRORQ\ 3&5 DPSOLILFDWLRQ IURP WKH 9) DQG 95 SULPHU DQQHDOLQJ VLWHV IODQNLQJ WKH %LR%ULFN LQVHUW RI S6%$ ZLWK ES DQG ES UHVSHF
WLYHO\ $JDURVH JHO HOHFWURSKRUHVLV RI DPSOLILHG '1$ UHYHDOHG UHOHYDQW EDQGV IRU DOO FORQHV LQGLFDWLQJ VXFFHVVIXO FORQLQJ RI WKH FRQVWUXFWHG SDUWV )LJXUH %%,QF4BUHS GDWD QRW VKRZQ $ ZHDN XQVSH
FLILF EDQG ZDV REVHUYHG LQ %%,QF4BUHS VDPSOHV DV WKLV ZDV DOVR REVHUYHG IRU WKH QHJDWLYH FRQWURO
WR ZKLFK QR WHPSODWH KDG EHHQ DGGHG WKLV ZDV GLVUHJDUGHG DV DQ DUWLIDFW IURP WKH 3&5 UHDFWLRQ
,QWHUHVWLQJO\ D VOLJKW GLIIHUHQFH LQ VL]H ZDV QRWHG IRU WKH IRXU %%,QF4BRUL9 FORQHV ZLWK & DQG SRVVLEO\ DOVR ' EHLQJ VRPHZKDW VPDOOHU WKDQ $ DQG % $ VHFRQG DJDURVH JHO GHGLFDWHG RQO\ WR WKH %%
,QF4BRUL9 VDPSOHV YHULILHG WKHVH REVHUYDWLRQV IRU FORQHV $ % DQG & ZLWK ' VWLOO VRPHZKDW XQFOHDU )LJXUH 7KLV VXJJHVWHG WKDW FORQHV $ DQG % FDUULHG WKH IXOOOHQJWK %%,QF4BRUL9 FRQVWUXFW ZKLOH FORQHV & DQG SRVVLEO\ ' SUREDEO\ FDUULHG WKH WUXQFDWHG YHUVLRQ
7ZR SRVLWLYH FORQHV RI HDFK FRQVWUXFW ZHUH VHQW IRU ILQDO YHULILFDWLRQ E\ VHTXHQFLQJ &ORQHV $ DQG % ZHUH VHTXHQFHG IRU DOO FRQVWUXFWV H[FHSW %%,QF4BUHS IURP ZKLFK FORQHV $ DQG & ZHUH VH
OHFWHG ,Q DGGLWLRQ DOO IRXU %%,QF4BRUL9 FORQHV $' ZHUH VHTXHQFHG WR LQFUHDVH WKH SUREDELOLW\ RI LGHQWLI\LQJ D IXOOOHQJWK SDUW $OO SDUWV ZHUH VHTXHQFHG ZLWK YHULILFDWLRQ SULPHUV 9) DQG 95 ZKLOH
%%,QF4BUHS ZDV DGGLWLRQDOO\ VHTXHQFHG ZLWK SULPHUV S6%; OLVWHG LQ WDEOH 8QIRUWXQDWHO\ UH
VXOWV IURP VHTXHQFLQJV GLG QRW DUULYH LQ WLPH WR EH LQFOXGHG LQ WKLV UHSRUW
)LJXUH $JDURVH JHO HOHFWURSKRUHVLV FDVH VWXG\ RI %%,QF4BRUL9 FORQHV$' 3UHYLRXVO\ DQDO\]HG FRORQ\ 3&5 SURGXFWVRIIRXU%%,QF4BRUL9 FORQHV ESZHUHUHDQDO\]HGRQD DJDURVHJHODW 9 PLQIRUDPRUH DFFXUDWHVHSDUDWLRQλ '1$ ODGGHU
9HULILFDWLRQ DQG FRQVWUXFWLRQ RI DQWLELRWLF UHVLVWDQFH %LR%ULFNV
9HULILFDWLRQ RI H[LVWLQJ %LR%ULFN DQWLELRWLF UHVLVWDQFH FDVVHWWHV
,Q RUGHU WR SUHSDUH D OLEUDU\ RI %LR%ULFN DQWLELRWLF UHVLVWDQFH PDUNHUV DYDLODEOH IRU XVH ZLWK S304$.
IRXU %LR%ULFN DQWLELRWLF UHVLVWDQFH FDVVHWWHV ZHUH H[WUDFWHG IURP '1$ UHSRVLWRU\ SODWHV REWDLQHG IURP WKH 5HJLVWU\ 7KH FDVVHWWHV HDFK FDUULHG E\ WKH S6%$ SODVPLG YHFWRU DQG UHVSHFWLYHO\ HQFRGLQJ UHVLVWDQFH WR DPSLFLOOLQ %%DB3 ES NDQDP\FLQ %%DB3 ES FKORUDPSKHQLFRO %%DB3 ES DQG WHWUDF\FOLQH %%DB3 ES ZHUH WHVWHG IRU H[SUHVVLRQ RI DQWLELRWLF UHVLVWDQFH LQ ( FROL JURZQ RYHUQLJKW ZLWK DSSURSULDWH DQWLELRWLFV $QWLELRWLF UHVLVWDQFH ZDV VXFFHVV
IXOO\ YHULILHG IRU %%DB3 %%DB3 DQG %%DB3 )RU %%DB3 QR FRQFOXVLRQ FRXOG EH GUDZQ DV DPSLFLOOLQ UHVLVWDQFH LV DOVR H[SUHVVHG IURP WKH S6%$ YHFWRU LWVHOI KRZHYHU VLQFH
%%DB3 LV D FRPPRQO\ XVHG UHVLVWDQFH FDVVHWWH DQG KDV SUHYLRXVO\ EHHQ VKRZQ WR H[SUHVV UHVLV
WDQFH WR DPSLFLOOLQ LW ZDV FRQFOXGHG WKDW YHULILFDWLRQ RI WKH FDVVHWWH¶V VL]H DQG QXFOHRWLGH VHTXHQFH VKRXOG EH VXIILFLHQW WR YHULI\ LWV IXQFWLRQ
7R FRQILUP WKH VL]H DQG SUHVHQFH RI DQWLELRWLF UHVLVWDQFH FDVVHWWHV LQ WKH FHOOV UHQGHUHG UHVLVWDQW
IRXU FORQHV $' IURP HDFK FRQVWUXFW ZHUH VHOHFWHG IRU YHULILFDWLRQ E\ FRORQ\ 3&5 IURP 9) DQG 95 SULPHU DQQHDOLQJ VLWHV IODQNLQJ WKH S6%$ %LR%ULFN LQVHUWV ZLWK ES DQG ES UHVSHFWLYHO\
$JDURVH JHO HOHFWURSKRUHVLV DQDO\VLV RI DPSOLILHG '1$ UHYHDOHG IUDJPHQWV ZLWK VL]HV FRUUHVSRQGLQJ DSSUR[LPDWHO\ WR WKH OHQJWKV H[SHFWHG IURP WKH VHTXHQFHV )LJXUH $OWKRXJK VPDOO GLIIHUHQFHV LQ VL]H ZHUH REVHUYHG EHWZHHQ FORQHV FDUU\LQJ WKH VDPH FRQVWUXFWV WKH VWHSZLVH LQFUHDVH DQG GHFUHDVH LQ VL]H RI WKHVH IUDJPHQWV VXJJHVWV D SUREOHP ZLWK WKH JHO UDWKHU WKDQ DFWXDO GLIIHUHQFHV LQ IUDJPHQW OHQJWKV
)RU ILQDO YHULILFDWLRQ WZR SRVLWLYH FORQHV $ DQG % RI HDFK FRQVWUXFW ZHUH VHQW IRU SDUWLDO VHTXHQF
LQJ ZLWK YHULILFDWLRQ SULPHUV 9) DQG 95 8QIRUWXQDWHO\ UHVXOWV IURP VHTXHQFLQJV GLG QRW DUULYH LQ WLPH WR EH LQFOXGHG LQ WKLV UHSRUW
&RQVWUXFWLRQ RI D QHZ VSHFWLQRP\FLQ UHVLVWDQFH %LR%ULFN IURP WKH S5/ SODVPLG )RU FRQVWUXFWLRQ RI D %LR%ULFN HQFRGLQJ UHVLVWDQFH WR VSHFWLQRP\FLQ %%6S5 ES WKH 7Q
WUDQVSRVRQ JHQH DDG$ FRGLQJ IRU VWUHSWRP\FLQ DGHQ\O\OWUDQVIHUDVH ZDV XVHG *HQ%DQN DFF QR
; $ 7QGHULYHG FRS\ RI WKLV JHQH ZDV IRXQG RQ SODVPLG S5/ DOVR H[SUHVVLQJ UH
VLVWDQFH WR DPSLFLOOLQ
)LJXUH &RORQ\ 3&5 YHULÀFDWLRQ RI %LR%ULFN DQWLELRWLF UHVLVWDQFH FDVVHWWHV )RXU FORQHV $' IURP HDFK RI WKH WHVWHG%LR%ULFNDQWLELRWLFUHVLVWDQFHFDVVHWWHV FDUULHGRQS6%$SODVPLGV ZHUHYHULÀHGE\FRORQ\3&5 IURP9)
DQG95 SULPHUDQQHDOLQJVLWHV ÁDQNLQJWKH%LR%ULFNLQVHUWVZLWK ES 3&5 SURGXFWVZHUHYHULÀHGIRUFRUUHFWVL]HV E\DJDURVHJHOHOHFWURSKRUHVLVRQ DJDURVHJHOVDW 9 IRU PLQ ([SHFWHGIUDJPHQWVL]HV %%DB3
ES %%DB3 ES %%DB3 ES %%DB3 ESλ 2ҋ*HQH5XOHUNE'1$ ODGGHU
([SUHVVLRQ RI VSHFWLQRP\FLQ UHVLVWDQFH IURP S5/ ZDV ILUVW WHVWHG DQG YHULILHG E\ WUDQVIRU
PDWLRQ LQWR ( FROL DQG JURZLQJ FHOOV RQ D VSHFWLQRP\FLQ SODWH 7KH %%6S5 %LR%ULFN ZDV WKHQ 3&5 DPSOLILHG IURP SUHSDUHG S5/ SODVPLG '1$ ZLWK SULPHUV LQWURGXFLQJ WKH %LR%ULFN SUHIL[ DQG VXIIL[ VHTXHQFHV DV SUHYLRXVO\ GHVFULEHG 7KH UHYHUVH SULPHU DOVR PRGLILHG WKH RULJLQDO VWRS FRGRQ WDD WR WZR VHTXHQWLDO VWRS FRGRQV WDDWDD LQ DJUHHPHQW ZLWK WKH %LR%ULFN VWDQGDUG $PSOLILHG '1$
ZDV DQDO\]HG E\ DJDURVH JHO HOHFWURSKRUHVLV ZKLFK UHYHDOHG D VLQJOH EDQG FRUUHVSRQGLQJ ZHOO WR WKH H[SHFWHG IUDJPHQW VL]H )LJXUH
$PSOLILHG %%6S5 '1$ ZDV GLJHVWHG ZLWK (FR5, DQG 3VW, IRU FORQLQJ LQWR SODVPLG YHFWRU S6%$
6LQFH ERWK WKH VRXUFH DQG WKH WDUJHW SODVPLG RI %%6S5 H[SUHVV UHVLVWDQFH WR DPSLFLOOLQ GLJHVWHG %%
6S5 '1$ ZDV VHSDUDWHG IURP S5/ SODVPLG E\ DJDURVH JHO HOHFWURSKRUHVLV DQG UHFRYHUHG E\ JHO H[WUDFWLRQ SULRU WR FORQLQJ 5HFRYHUHG %%6S5 '1$ ZDV WKHQ OLJDWHG LQWR S6%$ DQG YHFWRUV ZHUH WUDQVIRUPHG LQWR ( FROL DQG JURZQ RYHUQLJKW RQ DPSLFLOOLQ SODWHV WR VHOHFW IRU VXFFHVVIXOO\ WUDQVIRUPHG FORQHV )RXU FRORQLHV $' ZHUH WKHQ SLFNHG IRU FRORQ\ 3&5 YHULILFDWLRQ ZLWK 9) DQG 95 SULPHUV
$JDURVH JHO HOHFWURSKRUHVLV DQDO\VLV RI DPSOLILHG '1$ UHYHDOHG FOHDU VLQJOH EDQGV RI FRUUHFW VL]H IRU DOO IRXU FORQHV RI %%6S5 YHULI\LQJ VXFFHVVIXO FORQLQJ LQWR S6%$ )LJXUH
)LJXUH (OHFWURSKRUHVLV DQDO\VLV RI D FRQVWUXFWHG %%6S5 %LR%ULFN HQFRGLQJ UHVLVWDQFH WR VSHFWLQRP\FLQ 7KH
%LR%ULFNZDVÀUVW3&5 DPSOLÀHGIURPSODVPLGS5/ 6XFFHVVIXODPSOLÀFDWLRQZDVWKHQYHULÀHGE\DJDURVHJHO HOHFWURSKRUHVLV DJDURVH 9 PLQ OVDPSOH O2ҋ*HQH5XOHU ES'1$ ODGGHUλ ([SHFWHGSDUW VL]HLQFOXGHVVHTXHQFHVLQWURGXFHGE\ҋSULPHUH[WHQVLRQVDQGLVWKHUHIRUHORQJHUWKDQWKHDFWXDO%LR%ULFNSDUW
)LJXUH &RORQ\3&5 YHULÀFDWLRQRIFORQHG%%6S5 %LR%ULFN )RXUFORQHV$' RIWKHVXFFHVVIXOO\FORQHG%%
6S5 %LR%ULFN FDUULHGRQS6%$ ZHUHYHULÀHGE\FRORQ\3&5 IURP9)DQG95 SULPHUDQQHDOLQJVLWHV ÁDQNLQJWKH
%LR%ULFNLQVHUWZLWK ES 3&5 SURGXFWVZHUHYHULÀHGIRUFRUUHFWVL]HVE\DJDURVHJHOHOHFWURSKRUHVLVRQDQ DJDURVHJHODW 9 PLQ ([SHFWHGIUDJPHQWVL]H ESλ ES 2ҋ*HQH5XOHU ES'1$ ODGGHU
7ZR FORQHV % DQG ' ZHUH FKRVHQ IRU IXUWKHU FRQILUPDWLRQ DQG VHQW IRU VHTXHQFLQJ IURP 9)
DQG 95 SULPHU DQQHDOLQJ VLWHV 8QIRUWXQDWHO\ UHVXOWV IURP VHTXHQFLQJV GLG QRW DUULYH LQ WLPH WR EH LQFOXGHG LQ WKLV UHSRUW
'LVFXVVLRQ
:LWK WKH FRQVWUXFWLRQ DQG YHULILFDWLRQ RI QHZ DQG H[LVWLQJ %LR%ULFNV WKH EDVLF SDUWV IRU EXLOGLQJ D VL]HUHGXFHG YHUVLRQ RI S304$. DUH QRZ DYDLODEOH ,Q WKHLU DUWLFOH 6KHWW\ HW DO GHVFULEH D VWUDWHJ\ IRU DVVHPEO\ RI QHZ %LR%ULFN YHFWRUV XVLQJ D %LR%ULFN EDVH YHFWRU ,Q VKRUW WKH PHWKRG DOORZV IRU H[FKDQJH RI WKH EDVH YHFWRU¶V RULJLQDO UHSOLFRQ DQG DQWLELRWLF UHVLVWDQFH PDUNHU WR DQ\
QHZ FRPELQDWLRQ RI SODVPLG UHSOLFRQ DQG DQWLELRWLF UHVLVWDQFH FDVVHWWHV XVLQJ VWDQGDUG %LR%ULFN DVVHPEO\
)LJXUH LOOXVWUDWHV D SURSRVHG QHZ 56)GHULYHG YHFWRU S304.[ DVVHPEOHG IURP WKUHH RI P\ FRQVWUXFWHG SDUWV %%,QF4BRUL9 %%,QF4BUHS DQG %%53BRUL7 D NDQDP\FLQ UHVLVWDQFH FDV
VHWWH %%DB3 DQG WKUHH %LR%ULFNV UHDGLO\ DYDLODEOH IURP WKH 5HJLVWU\ DQG UHVSHFWLYHO\ FDUU\LQJ WKH %LR%ULFN EDVH YHFWRU %%DB, D ULERVRPDO ELQGLQJ VLWH 5%6 %%DB% IRU WUDQVODWLRQ RI 5HS%¶ DQG D FRQVWLWXWLYH SURPRWHU %%DB- IRU WUDQVFULSWLRQ RI WKH UHS%¶$& RSHURQ WKH ODW
WHU WZR ZHUH UDQGRPO\ FKRVHQ IRU LOOXVWUDWLRQ SXUSRVHV RQO\ $OVR %%DB3 ZDV LQFOXGHG LQ WKH YHFWRU DVVHPEO\ WR HQDEOH VL]H FRPSDULVRQ ZLWK WKH RULJLQDO S304$. YHFWRU
:LWK D VL]H RI RQO\ ES S304.[ PHDQV D UHGXFWLRQ LQ VL]H RI ES FRPSDUHG WR S304$. 7KLV UHGXFWLRQ LV LQ SDUW WKH UHVXOW RI UHPRYDO RI RQH RI WKH YHFWRU¶V DQWLELRWLF UHVLVWDQFH PDUNHUV EXW HYHQ ZKHQ WKH DPSLFLOOLQ UHVLVWDQFH FDVVHWWH %%DB3 LV LQFOXGHG WKH DVVHPEO\ OHDGV WR VL]H UHGXFWLRQ
,W LV RI FRXUVH QRW NQRZQ ZKHWKHU UHSODFLQJ WKH PRE UHJLRQ RI 56) ZLWK WKH 53RUL7 ZLOO UHVXOW LQ D PRELOL]DEOH SODVPLG WKLV ZLOO KDYH WR EH WHVWHG H[SHULPHQWDOO\ +RZHYHU RQH TXHVWLRQ RI FRQFHUQ WR EH DGGUHVVHG EHIRUH YHFWRU DVVHPEO\ LV WKH FKRLFH RI SURPRWHUV IRU UHS%¶$& WUDQVFULSWLRQ
$OWKRXJK D %LR%ULFN FDUU\LQJ WKH SS SURPRWHU FOXVWHU ZDV FRQVWUXFWHG WUDQVFULSWLRQ PD\ EH PRUH ILQHO\ WXQHG XVLQJ DOUHDG\ FKDUDFWHUL]HG SURPRWHUV IURP WKH 5HJLVWU\ $OVR DV WKH RULJLQDO S SURPRWHU SURPRWHV WUDQVFULSWLRQ LQ WKH RSSRVLWH GLUHFWLRQ 51$ SRO\PHUDVH ELQGLQJ WR S PD\ LQWHUIHUH ZLWK WUDQVFULSWLRQ IURP S DQG S FRPSOLFDWLQJ WUDQVFULSWLRQDO UHJXODWLRQ DQG WXQLQJ 7KLV PD\ EH DYRLGHG E\ FKRRVLQJ D VWDQGDORQH SURPRWHU
)LJXUH 3URSRVHGDVVHPEO\RIDVL]HUHGXFHGYHUVLRQRIWKHS304$.%LR%ULFNSODVPLGYHFWRU 7KHQHZYHFWRU ZDV DVVHPEOHG LQ VLOLFR DFFRUGLQJ WR WKH VWUDWHJ\ SURSRVHG E\ 6KHWW\ HW DO XVLQJ WKH %LR%ULFN EDVH YHF
WRU %%DB, WKUHH 56)GHULYHG %LR%ULFNV %%,QF4BRUL9 %%,QF4BUHS DQG %%53BRUL7 D NDQDP\FLQ UHVLVWDQFHFDVVHWWH%%DB3DQGWZRDGGLWLRQDO%LR%ULFNV%%DB%DQG%%DB-IURPWKH5HJLVWU\RI 6WDQGDUG%LRORJLFDO3DUWV 7KHUHVXOWLQJ ESYHFWRU JLYHQWKHZRUNLQJQDPHS304.[ LVDUHGXFWLRQRIWKHRULJL
QDOYHFWRUZLWK ES RU $EEUHYLDWLRQVGHVFULEHGLQERG\WH[W 3ODVPLGDVVHPEO\DQGLPDJHJHQHUDWHGZLWK
*HQHLRXV3URELRLQIRUPDWLFVVRIWZDUHKWWSZZZJHQHLRXVFRP
)XUWKHUPRUH WKUHH SURPRWHU UHJLRQV ORFDWHG LQ WKH 53RUL7 RI ZKLFK RQH SURPRWHV WUDQVFULSWLRQ LQ WKH GLUHFWLRQ RI UHS%¶$& 3DQVHJUDX HW DO PXVW DOVR EH WDNHQ LQWR FRQVLGHUDWLRQ DV WKH\
ZLOO DIIHFW WKH WRWDO WUDQVFULSWLRQ DFWLYLW\ 7KHVH SURPRWHU UHJLRQV PD\ SRVVLEO\ EH H[FOXGHG IURP WKH 53RUL7 RU LQDFWLYDWHG E\ PXWDJHQHVLV EXW VLQFH VXFK PRGLILFDWLRQV PD\ DOVR OHDG WR RWKHU FKDQJHV LQ WKH RUL7 IXQFWLRQDOLW\ WKLV ZLOO EH OHIW IRU IXWXUH ZRUN
0DWHULDOV DQG PHWKRGV
3UHSDUDWLRQ RI FRPSHWHQW FHOOV IRU WUDQVIRUPDWLRQ
&RPSHWHQW (VFKHULFKLD FROL '+α FHOOV ,QYLWURJHQ ZHUH SUHSDUHG E\ LQRFXODWLQJ PO VWHULOH O\VRJHQ\
EURWK /% JO %DFWRWU\SWRQH JO \HDVW H[WUDFW JO 1D&O ZLWK FHOOV IURP D ±
◦& JO\FHURO VWRFN DQG JURZQ DW
◦& ZLWK USP URWDU\ VKDNLQJ RYHUQLJKW PO IURP WKH RYHUQLJKW FXOWXUH ZDV WKHQ WUDQVIHUUHG WR D PO (IODVN ZLWK PO IUHVK /% DQG JURZQ DW
◦& ZLWK FRQWLQXRXV VKDNLQJ XQWLO D QP RSWLFDO GHQVLW\ 2'
600RI PHDVXUHPHQWV ZHUH EODQNHG ZLWK IUHVK /%
&HOOV ZHUH WUDQVIHUUHG WR [ PO FHQWULIXJH WXEHV DQG VSXQ GRZQ DW J DW
◦&IRU PLQ 6X
SHUQDWDQWV ZHUH GLVFDUGHG DQG SHOOHWV JHQWO\ UHVXVSHQGHG LQ PO LFH FROG &&0% EXIIHU P0 .2$F S+ P0 &D&O
2P0 0Q&O
2P0 0J&O
2JO\FHURO DQG LQFXEDWHG RQ LFH
PLQ &HQWULIXJDWLRQ ZDV UHSHDWHG ZLWK WKH VDPH VHWWLQJV DV EHIRUH DQG FHOOV ZHUH UHVXVSHQGHG LQ
PO QHZ &&0% EXIIHU DQG LQFXEDWHG PLQ RQ LFH &RPSHWHQW FHOOV ZHUH ILQDOO\ DOLTXRWHG DV
O VDPSOHV LQWR PO YLDOV DQG VWRUHG DW ±
◦&
7UDQVIRUPDWLRQV
$OO WUDQVIRUPDWLRQV ZHUH SHUIRUPHG ZLWK FRPSHWHQW (VFKHULFKLD FROL '+α FHOOV O FRPSHWHQW FHOOV WKDZHG IURP ±
◦& ZHUH PL[HG ZLWK O QJO SODVPLG '1$ DQG KHOG RQ LFH IRU PLQ
&HOOV ZHUH WKHQ KHDWVKRFNHG IRU V DW
◦& DQG LPPHGLDWHO\ UHWXUQHG WR LFH O /% ZDV DGGHG DQG FHOOV LQFXEDWHG PLQ DW
◦& ZLWK USP URWDU\ VKDNLQJ &HOOV ZHUH WKHQ VSXQ GRZQ DW IXOO VSHHG ≈ [ J IRU V DQG O ZDV UHPRYHG IURP WKH VXSHUQDWDQW &HOOV ZHUH ILQDOO\
UHVXVSHQGHG DQG SODWHG RQWR /% DJDU SODWHV J DJDUOLWHU ZLWK UHOHYDQW DQWLELRWLFV $PS $PS
&P $PS.P $PS6S $PS7F 7DEOH DQG JURZQ RYHUQLJKW DW
◦& WR VHOHFW IRU VXFFHVVIXOO\
WUDQVIRUPHG FHOOV
3ODVPLGV DQG %LR%ULFNV
([LVWLQJ %LR%ULFN SDUWV FDUULHG RQ %LR%ULFN SODVPLGV ZHUH REWDLQHG IURP WKH 5HJLVWU\ RI 6WDQGDUG
%LRORJLFDO 3DUWV KWWSSDUWVUHJLVWU\RUJ SURYLGHG RQ '1$ UHSRVLWRU\ SODWHV FRQWDLQLQJ GULHG '1$
'1$ ZDV UHVXVSHQGHG LQ O G+
22 IURP ZKLFK O ZDV WKHQ XVHG IRU WUDQVIRUPDWLRQ
3ODVPLG '1$ ZDV SUHSDUHG IURP RYHUQLJKW FHOO FXOWXUHV JURZQ LQ PO /% ZLWK DSSURSULDWH DQ
WLELRWLFV DW
◦& ZLWK USP URWDU\ VKDNLQJ XVLQJ WKH *HQ(OXWH 3ODVPLG 0LQLSUHS NLW IURP 6LJPD
$OGULFK IROORZLQJ WKH SURFHGXUHV LQ WKH SURYLGHG PDQXDO '1$ FRQFHQWUDWLRQV ZHUH WKHQ GHWHUPLQHG VSHFWURSKRWRPHWULFDOO\ DW QP ZLWK WKH /DPEHUW%HHU ODZ DQG VDPSOH SXULW\ SURWHLQ FRQWDPLQD
WLRQ ZDV FDOFXODWHG IURP WKH QP WR QP UDWLR ZLWK EDFNJURXQG FRUUHFWLRQ DW QP ,I QHFHVVDU\ VDPSOH FRQFHQWUDWLRQV ZHUH DGMXVWHG E\ GLOXWLRQ ZLWK G+
22 RU FRQFHQWUDWLRQ LQ DQ HYDSR
UDWLYH VDPSOH FRQFHQWUDWRU $OO SODVPLGV DQG %LR%ULFNV XVHG LQ WKH VWXG\ DUH OLVWHG LQ WDEOH
7DEOH &RQFHQWUDWLRQVRIDQWLELRWLFVIRUSUHSDUHG/% DJDUSODWHV
3ODWH &RQFHQWUDWLRQ RI DQWLELRWLF JPO
$PSLFLOOLQ &KORUDPSKHQLFRO .DQDP\FLQ 6SHFWLQRP\FLQ 7HWUDF\FOLQH
$PS
$PS&P
$PS.P
$PS6S
$PS7F
7DEOH 3ODVPLGVDQG%LR%ULFNVXVHGLQWKLVVWXG\
1DPH 'HVFULSWLRQ DQG XVDJH 5HIHUHQFH RU VRXUFH
S304$. %URDG KRVWUDQJH %LR%ULFN SODVPLG FORQLQJ YHFWRU %%,QF4B; %LR%ULFN DPSOLILFDWLRQ WHPSODWH
+XDQJ HW DO
S6%$ $PSLFLOOLQ UHVLVWDQFH KLJKFRS\ QXPEHU %LR%ULFN SODVPLG YHFWRU &ORQLQJ YHFWRU IRU FRQVWUXFWHG %LR%ULFNV
5HJLVWU\1
%%DB3 &\WRWR[LF SURWHLQ &FG%FRGLQJ %LR%ULFN &DUULHG E\ S6%$ 5HJLVWU\
S5/ 56) PRELOL]DWLRQ KHOSHU SODVPLG %%53BRUL7 %LR%ULFN DPSOLILFD
WLRQ WHPSODWH
(OKDL DQG :RON
S5/ 6SHFWLQRP\FLQ DQG VWUHSWRP\FLQ UHVLVWDQFH FORQLQJ YHFWRU %%6S5 %LR
%ULFN DPSOLILFDWLRQ WHPSODWH
& 3 :RON
%%DB3 $PSLFLOOLQ UHVLVWDQFH FDVVHWWH %LR%ULFN )XQFWLRQDO YHULILFDWLRQ DQG GHVLJQ RI VL]HUHGXFHG S304$. YHFWRU
5HJLVWU\
%%DB3 .DQDP\FLQ UHVLVWDQFH FDVVHWWH %LR%ULFN )XQFWLRQDO YHULILFDWLRQ DQG GHVLJQ RI VL]HUHGXFHG S304$. YHFWRU
5HJLVWU\
%%DB3 &KORUDPSKHQLFRO UHVLVWDQFH FDVVHWWH %LR%ULFN )XQFWLRQDO YHULILFDWLRQ 5HJLVWU\
%%DB3 7HWUDF\FOLQ UHVLVWDQFH FDVVHWWH %LR%ULFN )XQFWLRQDO YHULILFDWLRQ 5HJLVWU\
S6%$ $PSLFLOOLQ UHVLVWDQFH KLJKFRS\ QXPEHU %LR%ULFN SODVPLG YHFWRU &DUULHU RI
%%DB3
5HJLVWU\
%%DB% 5LERVRPDO ELQGLQJ VLWH 5%6 %LR%ULFN 'HVLJQ RI VL]HUHGXFHG S304$.
YHFWRU
%%DB- &RQVWLWXWLYH %LR%ULFN SURPRWHU 'HVLJQ RI VL]HUHGXFHG S304$. YHFWRU 5HJLVWU\
%%DB, %LR%ULFN EDVH YHFWRU 'HVLJQ RI VL]HUHGXFHG S304$. YHFWRU 5HJLVWU\
15HJLVWU\RI6WDQGDUG%LRORJLFDO3DUWVKWWSSDUWVUHJLVWU\RUJ
3ULPHUV
$OO SULPHUV XVHG LQ WKH VWXG\ DUH OLVWHG ZLWK VHTXHQFH DQG GHVFULSWLRQ LQ WDEOH 3ULPHUV XVHG IRU FRQ
VWUXFWLRQ RI %LR%ULFNV GHQRWHG %%SUH RU %%VXI ZHUH GHVLJQHG ZLWK VWDQGDUG %LR%ULFN SUHIL[ RU VXIIL[
VHTXHQFHV LQFOXGLQJ WKUHH DGGLWLRQDO ¶ EDVHV *$$ WR IDFLOLWDWH HQGRQXFOHDVH GLJHVWLRQ SUHIL[ VH
TXHQFH YDULHG GHSHQGLQJ RQ ZKHWKHU LW ZDV IROORZHG E\ D SURWHLQFRGLQJ RU QRQFRGLQJ SDUW 3ULPHUV ZHUH SXUFKDVHG IURP 7KHUPR )LVKHU 6FLHQWLILF *PE+ 3ULPHU DOLJQPHQWV ZHUH FDUULHG RXW XVLQJ WKH
%DVLF /RFDO $OLJQPHQW 6HDUFK 7RRO %/$67 SURYLGHG RQOLQH E\ WKH 1DWLRQDO &HQWHU IRU %LRWHFKQRO
RJ\ ,QIRUPDWLRQ 1&%, KWWSEODVWQFELQOPQLKJRY%ODVWFJL 9HULILFDWLRQ SULPHUV 9) DQG 95
DV ZHOO DV VHTXHQFLQJ SULPHUV S6%; ZHUH NLQGO\ SURYLGHG E\ 'DQLHO &DPVXQG 8SSVDOD 8QLYHUVLW\
'1$ VHTXHQFLQJ
3ODVPLG '1$ ZDV VHQW IRU '1$ VHTXHQFLQJ WR 0DFURJHQ ,QF
&RQVWUXFWLRQ DQG FORQLQJ RI QHZ %LR%ULFN SDUWV
$OO FRQVWUXFWHG %LR%ULFN SDUWV ZHUH DPSOLILHG E\ 3&5 XVLQJ WKH 3KXVLRQ +LJK)LGHOLW\ '1$ 3RO\
PHUDVH IURP )LQQ]\PHV 3&5 UHDFWLRQ WXEHV ZHUH SUHSDUHG ZLWK O P0 [ 3KXVLRQ +) EXIIHU
O P0 G173V O 0 IRUZDUG DQG UHYHUVH SULPHUV O P0 3KXVLRQ '1$ SRO\
PHUDVH O 0 WHPSODWH SODVPLG DQG G+
22 WR D WRWDO YROXPH RI O 3URGXFWV ZHUH WKHQ 3&5 DPSOLILHG ZLWK F\FOHV RI V GHQDWXUDWLRQ DW
◦& V SULPHU DQQHDOLQJ DW
◦& DQG HORQJDWLRQ DW
◦& IROORZHG E\ F\FOHV RI V GHQDWXUDWLRQ DW
◦& V SULPHU DQQHDOLQJ DW
◦& DQG HORQ
JDWLRQ DW
◦& (ORQJDWLRQ WLPHV ZHUH VHW WR V IRU DPSOLILFDWLRQ RI %%,QF4B3 %%,QF4BRUL9
%%,QF4BRUL7 %%,QF4B33 %%53BRUL7 DQG %%6S5 PLQ V IRU %%,QF4B56) DQG
%%,QF4BUHS $GGLWLRQDO V GHQDWXUDWLRQ DQG PLQ HORQJDWLRQ VWHSV ZHUH LQFOXGHG EHIRUH DQG DIWHU WKH DPSOLILFDWLRQ F\FOHV UHVSHFWLYHO\
6XFFHVVIXO 3&5 DPSOLILFDWLRQ ZDV YHULILHG E\ DJDURVH JHO HOHFWURSKRUHVLV $JDURVH JHOV ZHUH FDVW E\ PHOWLQJ DJDURVH LQ VRGLXP ERUDWH 6% EXIIHU P0 1D2+ +
3%2
3WR D S+ RI FRQWDLQLQJ
O WKLD]ROH RUDQJH '1$ VWDLQLQJ DJHQW IURP D VWRFN PJ SRZGHU LQ PO '062 SHU OLWHU 6%
WR \LHOG D ILQDO DJDURVH FRQFHQWUDWLRQ RI GHSHQGLQJ RQ WKH VL]H RI WKH '1$ WR EH VHSDUDWHG ! NE NE NE O VDPSOHV ZHUH ORDGHG ZLWK O ; JHO ORDGLQJ G\H JO\FHURO EURPRSKHQRO EOXH DQG JHOV UXQ LQ 6% UXQQLQJ EXIIHU IRU PLQ DW
9 GHSHQGLQJ RQ '1$ IUDJPHQWV JHO FRQFHQWUDWLRQ DQG GHVLUHG VHSDUDWLRQ SUHFLVLRQ
9HULILHG %LR%ULFNV ZHUH SUHSDUHG IRU FORQLQJ LQWR S6%$ %LR%ULFN SODVPLG YHFWRUV E\ GLJHVWLQJ DPSOLILHG '1$ DQG S6%$ SODVPLGV FDUU\LQJ WKH %%DB3 %LR%ULFN FRGLQJ IRU WKH F\WRWR[LQ SURWHLQ &FG% ZLWK )DVW'LJHVW UHVWULFWLRQ HQ]\PHV IURP )HUPHQWDV 'LJHVWLRQ WXEHV ZHUH SUHSDUHG ZLWK O ; )DVW'LJHVW EXIIHU O J SODVPLG '1$ RU O 3&5 SURGXFW O )DVW'LJHVW (FR5, O )DVW'LJHVW 3VW, DQG G+
22 WR D WRWDO YROXPH RI O 'LJHVWLRQ ZDV FDUULHG RXW IRU
PLQ at
◦C DQG FRPSOHWH GLJHVWLRQ YHULILHG RQ DJDURVH JHOV 'LJHVWHG DPSOLILHG '1$
DQG OLQHDUL]HG S6%$ SODVPLG ZHUH WKHQ VHSDUDWHG IURP WHPSODWH SODVPLG DQG H[FLVHG %%DB3
LQVHUWV E\ DJDURVH JHO HOHFWURSKRUHVLV 5HOHYDQW '1$ ZDV UHFRYHUHG E\ H[FLVLQJ EDQGV IURP WKH JHOV DQG SXULI\LQJ '1$ XVLQJ WKH 1XFOHR6SLQ ([WUDFW ,, NLW IURP 0DFKHUH\1DJHO DFFRUGLQJ WKH WKH PDQXIDFWXUHU¶V LQVWUXFWLRQV 'LJHVWHG SDUWV ZHUH WKHQ OLJDWHG LQWR S6%$ DW LQVHUWWRYHFWRU UDWLRV XVLQJ WKH 4XLFN /LJDWLRQ NLW IURP 1HZ (QJODQG %LRODEV DFFRUGLQJ WR WKH SURYLGHG LQVWUXFWLRQV
$IWHU OLJDWLRQ YHFWRUV ZHUH WUDQVIRUPHG LQWR FKHPLFDOO\ FRPSHWHQW FHOOV IRU FORQLQJ
6XFFHVVIXOO\ WUDQVIRUPHG FORQHV ZHUH DQDO\]HG IRU LQVHUW OHQJWK E\ FRORQ\ 3&5 &RORQ\ 3&5 DPSOLILFDWLRQV ZHUH SHUIRUPHG ZLWK WKH 'UHDP7DT '1$ 3RO\PHUDVH IURP )HUPHQWDV 3&5 WHPSODWHV ZHUH SUHSDUHG E\ SLFNLQJ VLQJOH FRORQLHV UHSUHVHQWLQJ VXFFHVVIXOO\ WUDQVIRUPHG FHOOV IURP /% DJDU SODWHV DQG UHVXVSHQGLQJ HDFK FRORQ\ LQ O /% 3&5 UHDFWLRQ WXEHV ZHUH SUHSDUHG ZLWK O P0
; 'UHDP7DT EXIIHU O P0 G173V O 0 IRUZDUG 9) DQG UHYHUVH 95 SULPHUV
O P0 'UHDP7DT '1$ SRO\PHUDVH O WHPSODWH FHOO VXVSHQVLRQ DQG G+
22 WR D WRWDO YROXPH RI O 3URGXFWV ZHUH WKHQ 3&5 DPSOLILHG ZLWK F\FOHV RI V GHQDWXUDWLRQ DW
◦&
V SULPHU DQQHDOLQJ DW
◦& DQG HORQJDWLRQ DW
◦& HORQJDWLRQ WLPHV ZHUH VHW WR PLQ V IRU
%%,QF4BUHS DQG PLQ V IRU %%,QF4BRUL9 %%,QF4BRUL7 %%,QF4B33 %%6S5 DQG %%
53BRUL7 $GGLWLRQDO PLQ GHQDWXUDWLRQ DQG HORQJDWLRQ VWHSV ZHUH LQFOXGHG EHIRUH DQG DIWHU WKH
F\FOHV UHVSHFWLYHO\ 3&5 SURGXFWV ZHUH DQDO\]HG E\ DJDURVH JHO HOHFWURSKRUHVLV DV GHVFULEHG DERYH
9HULILHG FORQHV ZHUH VHOHFWHG IRU SODVPLG '1$ SUHSDUDWLRQ DQG '1$ VHTXHQFLQJ DQG JO\FHURO VWRFNV ZHUH SUHSDUHG DQG VWRUHG LQ ±
◦&
9HULILFDWLRQ RI H[LVWLQJ DQWLELRWLF UHVLVWDQFH FDVVHWWH %LR%ULFNV
([LVWLQJ %LR%ULFNV ZHUH REWDLQHG IURP WKH 5HJLVWU\ RI 6WDQGDUG %LRORJLFDO 3DUWV KWWSSDUWVUHJLVWU\
RUJ RQ '1$ UHSRVLWRU\ SODWHV DV GULHG '1$ '1$ ZDV H[WUDFWHG IURP SODWHV E\ UHVXVSHQGLQJ LQ
O G+
22 IURP ZKLFK O ZDV H[WUDFWHG IRU WUDQVIRUPDWLRQ 7UDQVIRUPHG FHOOV ZHUH JURZQ RQ /% DJDU SODWHV ZLWK UHOHYDQW DQWLELRWLFV DFFRUGLQJ WR SODVPLG DQG FDVVHWWH UHVLVWDQFH 6XFFHVVIXOO\
WUDQVIRUPHG FORQHV ZHUH DQDO\]HG IRU LQVHUW OHQJWK E\ FRORQ\ 3&5 DV GHVFULEHG DERYH ZLWK HORQJD
WLRQ WLPHV VHW WR PLQ V 9HULILHG FORQHV ZHUH VHOHFWHG IRU SODVPLG '1$ SUHSDUDWLRQ DQG '1$
VHTXHQFLQJ DQG JO\FHURO VWRFNV ZHUH SUHSDUHG DQG VWRUHG LQ ±
◦&
7DEOH3ULPHUVXVHGIRU3&5DPSOLÀ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À[RUVXIÀ[VHTXHQFHV