|
|
 |
| l |
¤°»ò¬O®Ö»Ä©w§Ç (DNA Sequencing)? |
| |
«ü¤ÀªR¯S©wDNA¤ù¬qªºÆP°ò(DNA¤À¤lªº°ò¥»ºc³y³æ¦ì)§Ç¦C¡A¤]´N¬O¸¢áIËï(A)¡B¯Ý¸¢áGÔr(T)¡BMáGÔr(C)»P³¾áIËï(G)±Æ¦C¤è¦¡¡C
RNA©w§Ç«h³q±`±NRNA´£¨ú«á¡A¤ÏÂà¿ý¬°DNA«á¨Ï¥ÎDNA©w§Çªº¤èªk¶i¦æ©w§Ç¡C
¥Ø«eÀ³¥Î³Ì¼sªxªº¬O¥ÑFrederick
Sangerµo©úªºSangerÂù²æ®ñÃì²×¤îªk(Chain
Termination Method)¡C·sªº©w§Ç¤èªk¡A¦³µJÁC»Ä©w§Çªk©M454©w§Çªk¡C
|
| 2. |
¬°¤°»òn©e¥~¥N¤u? |
| |
¹ï¤À¤l¥Íª«¬ã¨sªÌ¦Ó¨¥, ®Ö»Ä©w§Ç¼Æ¾Ú¬O·¥¬°°ò¥»¥B«nªº¸ê®Æ¡C¦Ón¦p¦ó±o¨ì¥¿½T¦Ó¥i¾aªº©w§Ç¸ê®Æ,
«o¬OÓ¥O¤HÀY¯kªº°ÝÃD¡C¥ý¤£½×°Ê»³¼Æ¦Ê¬Æ¦Ü¤W¤d¸Uªº©w§Ç»ö¦p¦ó¨ú±o, §Y¨Ï¦³»ö¾¹¥i¥H¨Ï¥Î, n«Ø¥ß¤@Ó¨}¦nªº©w§Ç¹êÅç¨t²Î¤]¬O·¥¨ä¯Ó¶O®É¶¡¤Î¬ã¨s¸ê·½ªº¨Æ¡CÁöµM²{¦b¤w¦³¼Æ®a¥N«È©w§Ç¤½¥q¥i´£¨Ñ¬ÛÃöªA°È,
¦ý¬O°Ñ®t¤£»ôªºªA°È«~½è, «o±`Åý¤H´£¤ß¦QÁx¡K ¤ñ¸ûÂÔ·Vªº¹êÅç«Ç, §ó±`±`¦P®É±N¼Ë«~©e°U¤£¦Pªº¤½¥q¶i¦æ©w§Ç, ¥H½T»{¼Æ¾Úªº¥¿½T©Ê;
¦ý¬O¦p¦¹«o¤S³y¦¨¸ê·½ªº®ö¶O¡C§Ú̧Ʊæ¯à´£¨Ñ±z¤@Ó¤£¤@¼Ëªº¿ï¾Ü, Åý±z¯à°÷¦w¤ß, ¦³®Ä²vªº±o¨ì¥¿½Tªº©w§Ç¼Æ¾Ú¡C |
| |
|
| 3. |
§Ú̪º¯SÂI |
| |
| A. |
«~½èéw
| ©w§Çªø«×¥§¡¥i¹F650bp¥H¤W,
¥¿½T²v§ó¥i¹F99.5% (¦³¹Ï¬°ÃÒ)¡Cn¦p¦ó§PÂ_®Ö»Ä©w§Ç¼Æ¾Úªº¦nÃa©O? ³oùØ´£¨Ñ±z´XÂI§@°Ñ¦Ò: |
| 1.
|
I´ºÈªº°ª§C: ¦³ªº¦Ñ®v¦b±q¨Æªº¬ã¨s´N¬OÂI¬ðÅÜ
(Point Mutation) ©Î¬O³æ®Ö?¦h«¬©Ê (Single Nucleotide Polymorphism,
SNPs) ¤ÀªR , ¦ý¬O¨¯¨¯WW Construct ¦nªº Plasmid ©Î¬O¦n¤£®e©ö§¨¥X¨Óªº
PCR products °e¥X¥h©w§Ç, ¦^¨Ó®É«oµLªk§P©wµ²ªG¬ODouble peaks ÁÙ¬O Background
¡C¦]¦¹¤@¥÷¨}¦nªº®Ö»Ä©w§Ç¼Æ¾ÚÀ³¸Ó¨ã¦³«Ü§CªºI´ºÈ, ¤~¯à¼W¥[§PŪªº®e©ö«×¡C |
| 2.
|
¦Û¥Ñ¬V¾¯ (Free Dye)
ªº¦h¹è: ©Ò¿×ªºFree Dye ´N¬O«Üµuªº Flourescent labeled ddNTP,
¤]´N¬O¦U¦ì±`¬Ý¨ì¦ì©ó0-30bp ¥ª¥k¤@³s¦ê«Ü°ªªºpeak. |
| 3. |
®p«¬ (Peak) ªº¦y¾U«×
(Sharpness) |
| 4.
|
©w§Çªø«× |
| 5.
|
¨ä¥L |
|
| B. |
»ù®æ¦X²z
µL½×±zªº¼Ë«~¬Oµß®è, ½èÅé§í©Î¬OPCR ²£ª«, §Ṳ́@«ß³£¥u¦¬NTD 450/sample, °w¹ï¶q¤j©Î¦³¯S®í»Ý¨Dªº«È¤á,
¥t¦³p»ù¤è¦¡, Åwªï¨Ó¹q¬¢¸ß¡C |
| C. |
ªA°È¿Ë¤Á
¤j¥x¥_¦a°Ï¦³±M¤H¦¬°e¥ó, ¦¬¥ó«á¤T¤é¤º§Y¥i±o¨ìµ²ªG¡C¥~¿¤¥«¦b¼Ë«~©è¹F«á¤T¤é¤º¥ç¥i±o¨ìµ²ªG¡C |
| D. |
§Þ³N±M·~
¤£¹³¨ä¥¦ªº©w§ÇªA°È¤½¥q, §Ú̬ñ¤¨C¦ì«È¤áªº©e°U¡C¦]¬°§Ú̪¾¹D, °e¨ì§Ú̹êÅç«Çªº¨CӼ˫~, ³£¬O±z¤ß¦åªºµ²´¹¡C¤]¦]¦¹,
¨CÓ©w§Çµ²ªG,³£·|¸g¹L§Ú̪º¦A¤T½T»{¤§«á¤~·|°e¥X¥h¡CY¬O¦b©w§Çªº¹Lµ{¤¤¥X²{¤F°ÝÃD, §Ṳ́]·|¥ý¥Î¹q¸Ü©Î¹q¤l¶l¥ó»P±z½T»{¡C(©w§Ç¥¢±Ñ¼Ë«~¦¬¶O¼Ð·Ç) |
|
| |
|
| |
|
|
| |
|