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SILVER ๋“ฑ๊ธ‰์˜ ํŒ๋งค์ž ์ž๋ฃŒ

T-vector์™ฟ’ insert DNA ๋น„์œจ์„ ๋‹ฌ๋ฆฌํ•˜์—ฌ TA cloning์„ ํ†ตํ•ด ์–ป์–ด์ง„ colony ๋น„๊ต ๋ถ„์„

"T-vector์™ฟ’ insert DNA ๋น„์œจ์„ ๋‹ฌ๋ฆฌํ•˜์—ฌ TA cloning์„ ํ†ตํ•ด ์–ป์–ด์ง„ colony ๋น„๊ต ๋ถ„์„"์— ๋Œ€ํ•œ ๋‚ด์šฉ์ž…๋‹ˆ๋‹ค.
8 ํŽ˜์ด์งฟ’
์›Œ๋“œ
์ตœ์ดˆ๋“ฑ๋ก์ผ 2020.09.22 ์ตœ์ข…์ ฟ’์ž‘์ผ 2020.09
8P ๋ฏธ๋้ฉ๋ณด๊ธฐ
T-vector์™ฟ’ insert DNA ๋น„์œจ์„ ๋‹ฌ๋ฆฌํ•˜์—ฌ TA cloning์„ ํ†ตํ•ด ์–ป์–ด์ง„ colony ๋น„๊ต ๋ถ„์„
  • ๋ฏธ๋้ฉ๋ณด๊ธฐ

    ์†Œ๊ฐœ

    "T-vector์™ฟ’ insert DNA ๋น„์œจ์„ ๋‹ฌ๋ฆฌํ•˜์—ฌ TA cloning์„ ํ†ตํ•ด ์–ป์–ด์ง„ colony ๋น„๊ต ๋ถ„์„"์— ๋Œ€ํ•œ ๋‚ด์šฉ์ž…๋‹ˆ๋‹ค.

    ๋ชฉ์ฐจ

    1. Subject
    2. Object
    3. Introduction
    4. Material & Method
    5. Result
    6. Discussion

    ๋ณธ๋ๅฉ๋‚ด์šฉ

    1. Subject : recombinant plasmid DNA cloning

    2. Object: T-vector์™ฟ’ insert DNA ๋น„์œจ์„ ๋‹ฌ๋ฆฌํ•˜์—ฌ TA cloning์„ ํ†ตํ•ด ์–ป์–ด์ง„ colony๋ฅผ ๋น„๊ต ๋ถ„์„ํ•ด๋ณธ๋‹ค.

    3. Introduction
    TA cloning : ๋ณดํ†ต PCR product๋Š” 3โ€™ ๋ง๋‹จ์ด blunt end๋กœ ๋˜์–ด ์žˆ๋Š” ์ค„ ์•Œ์ง€๋งŒ ๊ทธ๋ ‡์ง€ ์•Š์€ ๊ฒฝ์šฐ๊ฐ€ ๋งŽ๋‹ค. ํŠนํžˆ PCR์— ์‚ฌ์šฉํ•˜๋Š” polymerase๋Š” Thermus aquaticus (Taq) polymerase๋ฅผ ๋งŽ์ด ์‚ฌ์šฉํ•˜๋Š”๋ฐ, Taq polymerase๋Š” Terminal deoxynucleotidyl transferase๋กœ์„œ Extension์ด ๋๋‚ ๋•Œ 3โ€™๋ง๋‹จ์— nucleotide A๋ฅผ ๋” ๋ถ™์ด๊ณ  ๋์ด ๋‚˜๋Š” ํŠน์„ฑ์„ ๊ฐ€์ง€๊ณ  ์žˆ๋‹ค. ๋‹จ ๋‹จ์ง€ ํ•œ ๊ฐœ์˜ A๋งŒ์„ ๋ถ™์ด๊ณ  ๋์ด ๋‚˜๊ธฐ ๋•Œ๋ฌธ์— DNA๋ฅผ cloning์„ ํ•˜๊ณ  ์‹ถ๋‹ค๋ฉด vector์˜ 5โ€™ ๋ง๋‹จ์ด T๋กœ ๋๋‚˜๋Š” vector๋ฅผ ์‚ฌ์šฉํ•˜๋ฉด ๋˜๊ณ  ๊ทธ๋ ‡๊ธฐ์— ์ด cloning์ด TA cloning์ด๋ผ๊ณ  ๋ถˆ๋ฆฌ๋Š” ๊ฒƒ์ด๋‹ค.
    T-vector๋ฅผ ๋งŒ๋“œ๋Š” ๋ฐฉ๋ฒ•์€ ์–ด๋– ํ•œ vector์— ์ƒ๊ด€์—†์ด blunt end๋ฅผ ๋งŒ๋“œ๋Š” ์ œํ•œํšจ์†Œ๋ฅผ ์ฒ˜๋ฆฌํ•œ ํ›„ ์ „๊ธฐ์˜๋™์„ ํ•œ ํ›„ gel purifiction์„ ํ•˜๊ณ  ์ด product 5ul์— dTTP 2mM(final), 10X Taq buffer, Taq polymerase(5unit)๋ฅผ ๋„ฃ๊ณ  70โ„ƒ 2์‹œ๊ฐ„ PCRํ•œ๋‹ค์Œ PCR product๋ฅผ ์ •์ œํ•œ ํ›„ ์‚ฌ์šฉํ•œ๋‹ค๊ณ  ํ•œ๋‹ค.
    ์ด๋Ÿฌํ•œ T-vector๋ฅผ ์ด์šฉํ•˜๋Š” ์ด์œ ๋Š” ์—ฌ๋Ÿฌ๊ฐ€์ง€๊ฐ€ ์žˆ์„ ์ˆ˜ ์žˆ๋Š”๋ฐ ์šฐ์„  PCR product์–‘์ด ๋„ˆ๋ฌด ์ ์„ ๊ฒฝ์šฐ cloning์„ ์œ„ํ•œ PCR๋ฐ˜์‘ ์‚ฐ๋ฌผ์ด ๋„ˆ๋ฌด ์ ์„ ๊ฒฝ์šฐ ์ œํ•œํšจ์†Œ ์ฒ˜๋ฆฌ ๋ฐ DNA์ถ”์ถœ ๊ณผ์ •์— ๋”ฐ๋ฅธ insert DNA ๋†๋„๊ฐ€ ์ค„๊ฒŒ ๋จ์— ๋”ฐ๋ผ vector์— ์‚ฝ์ž… ํ• ๋•Œ ๋ถ€๋‹ด์ด ํฌ๋‹ค. ๊ทธ๋ ‡๋‹ค๊ณ  PCR reaction cycle์ˆ˜๋ฅผ ๋Š˜๋ฆฌ๋ฉด ์ค‘๊ฐ„์— mutation์ด ๋ฐœ์ƒํ•  ์šฐ๋ ค๊ฐ€ ์žˆ๋‹ค.

    ์ฐธ๊ณ ์ž๋ฃŒ

    ยท Zhou,M.Y., Clark,S.E. and Gomez-Sanchez, C.E.(1995)
    ยท Universal cloning method by TA strategy. BioTechniques 19,34.
    ยท https://m.ibric.org/miniboard/read.php?Board=exp_qna&id=83556
    ยท https://www.sciencedirect.com/science/article/abs/pii/S0168165602001074
  • ์ž๋ฃŒํ›„๊ธฐ

      Ai ๋ฆฌ๋ทฐ
      ์งฟ’์‹ํŒ๋งค์ž๊ฐ€ ๋“ฑ๋กํ•œ ์ž๋ฃŒ๋Š” ๊ณผ์ œ์— ์ ์šฉํ•  ์ˆ˜ ์žˆ๋Š” ์œ ์šฉํ•œ ๋‚ด์šฉ์ด ๋งŽ์•„, ํฐ ๋„์›€์ด ๋˜์—ˆ์Šต๋‹ˆ๋‹ค. ์•ž์œผ๋กœ๋„ ๋งŽ์€ ๋„์›€์„ ๋ฐ›์„ ์ˆ˜ ์žˆ๊ธฐ๋ฅผ ๊ธฐ๋Œ€ํ•ฉ๋‹ˆ๋‹ค!
    • ์ž์ฃผ๋ฌป๋Š”์งˆ๋ๅฉ์˜ ๋‹ต๋ณ€์„ ํ™•์ธํ•ด ์ฃผ์„ธ์š”

      ํ•ดํ”ผ์บ ํผ์Šค FAQ ๋”๋ต–๊ธฐ

      ๊ผญ ์•Œ์•„์ฃผ์„ธ์š”

      • ์ž๋ฃŒ์˜ ์ •๋ณด ๋ฐ ๋‚ด์šฉ์˜ ์ง„์‹ค์„ฑ์— ๋Œ€ํ•˜์—ฌ ํ•ดํ”ผ์บ ํผ์Šค๋Š” ๋ณด์ฆํ•˜์ง€ ์•Š์œผ๋ฉฐ, ํ•ด๋‹น ์ •๋ณด ๋ฐ ๊ฒŒ์‹œ๋ฌผ ์ €์ž‘๊ถŒ๊ณผ ๊ธฐํƒ€ ๋ฒ•์  ์ฑ…์ž„์€ ์ž๋ฃŒ ๋“ฑ๋ก์ž์—๊ฒŒ ์žˆ์Šต๋‹ˆ๋‹ค.
        ์ž๋ฃŒ ๋ฐ ๊ฒŒ์‹œ๋ฌผ ๋‚ด์šฉ์˜ ๋ถˆ๋ฒ•์  ์ด์šฉ, ๋ฌด๋‹จ ์ „์žฌโˆ™๋ฐฐํฌ๋Š” ๊ธˆ์ง€๋˜์–ด ์žˆ์Šต๋‹ˆ๋‹ค.
        ์ €์ž‘๊ถŒ์นจํ•ด, ๋ช…์˜ˆํ›ผ์† ๋“ฑ ๋ถ„์Ÿ ์š”์†Œ ๋ฐœ๊ฒฌ ์‹œ ๊ณ ๊ฐ๋น„๋ฐ”์นด์ง€๋…ธ Viva์˜ ์ €์ž‘๊ถŒ์นจํ•ด ์‹ ๊ณ ๋น„๋ฐ”์นด์ง€๋…ธ Viva๋ฅผ ์ด์šฉํ•ด ์ฃผ์‹œ๊ธฐ ๋ฐ”๋ž๋‹ˆ๋‹ค.
      • ํ•ดํ”ผ์บ ํผ์Šค๋Š” ๊ตฌ๋งค์ž์™ฟ’ ํŒ๋งค์ž ๋ชจ๋‘๊ฐ€ ๋งŒ์กฑํ•˜๋Š” ์„œ๋น„์Šค๊ฐ€ ๋˜๋„๋ก ๋…ธ๋ ฅํ•˜๊ณ  ์žˆ์œผ๋ฉฐ, ์•„๋ž˜์˜ 4๊ฐ€์ง€ ์ž๋ฃŒํ™˜๋ถˆ ์กฐ๊ฑด์„ ๊ผญ ํ™•์ธํ•ด์ฃผ์‹œ๊ธฐ ๋ฐ”๋ž๋‹ˆ๋‹ค.
        ํŒŒ์ผ์˜ค๋ฅ˜ ์ค‘๋ณต์ž๋ฃŒ ์ €์ž‘๊ถŒ ์—†์Œ ์„ค๋ช…๊ณผ ์‹ค์ œ ๋‚ด์šฉ ๋ถˆ์ผ์น˜
        ํŒŒ์ผ์˜ ๋‹ค์šด๋กœ๋“œ๊ฐ€ ์ œ๋Œ€๋กœ ๋˜์ง€ ์•Š๊ฑฐ๋‚˜ ํŒŒ์ผํ˜•์‹์— ๋งž๋Š” ํ”„๋กœ๊ทธ๋žจ์œผ๋กœ ์ •์ƒ ์ž‘๋™ํ•˜์ง€ ์•Š๋Š” ๊ฒฝ์šฐ ๋‹ค๋ฅธ ์ž๋ฃŒ์™ฟ’ 70% ์ด์ƒ ๋‚ด์šฉ์ด ์ผ์น˜ํ•˜๋Š” ๊ฒฝ์šฐ (์ค‘๋ณต์ž„์„ ํ™•์ธํ•  ์ˆ˜ ์žˆ๋Š” ๊ทผ๊ฑฐ ํ•„์š”ํ•จ) ์ธํ„ฐ๋„ท์˜ ๋‹ค๋ฅธ ์‚ฌ์ดํŠธ, ์—ฐ๊ตฌ๊ธฐ๊ด€, ํ•™๊ป“, ์„œ์  ๋“ฑ์˜ ์ž๋ฃŒ๋ฅผ ๋„์šฉํ•œ ๊ฒฝ์šฐ ์ž๋ฃŒ์˜ ์„ค๋ช…๊ณผ ์‹ค์ œ ์ž๋ฃŒ์˜ ๋‚ด์šฉ์ด ์ผ์น˜ํ•˜์ง€ ์•Š๋Š” ๊ฒฝ์šฐ

    ์ฐพ์œผ์‹œ๋˜ ์ž๋ฃŒ๊ฐ€ ์•„๋‹Œ๊ฐ€์š”?

    ์ง€๊ธˆ ๋ณด๋Š” ์ž๋ฃŒ์™ฟ’ ์—ฐ๊ด€๋˜์–ด ์žˆ์–ด์š”!
    ์™ผ์ชฝ ํ™”์‚ดํ‘œ
    ์˜ค๋ฅธ์ชฝ ํ™”์‚ดํ‘œ
    ๋ฌธ์„œ ์ดˆ์•ˆ์„ ์ƒ์„ฑํ•ด์ฃผ๋Š” EasyAI
    ์•ˆ๋…•ํ•˜์„ธ์š”. ํ•ดํ”ผ์บ ํผ์Šค์˜ ๋ฐฉ๋Œ€ํ•œ ์ž๋ฃŒ ์ค‘์—์„œ ์„ ๋ณ„ํ•˜์—ฌ ๋‹น์‹ ๋งŒ์˜ ์ดˆ์•ˆ์„ ๋งŒ๋“ค์–ด์ฃผ๋Š” EasyAI ์ž…๋‹ˆ๋‹ค.
    ์ €๋Š” ์•„๋ž˜์™ฟ’ ๊ฐ™์ด ์ž‘์—…์„ ๋„์™ฟ’๋“œ๋ฆฝ๋‹ˆ๋‹ค.
    - ์ฃผ์ œ๋งŒ ์ž…๋ ฅํ•˜๋ฉด ๋ชฉ์ฐจ๋ถ€ํ„ฐ ๋ณธ๋ๅฉ๋‚ด์šฉ๊นŒ์ง€ ์ž๋™ ์ƒ์„ฑํ•ด ๋“œ๋ฆฝ๋‹ˆ๋‹ค.
    - ์žฅ๋ฌธ์˜ ์ฝ˜ํ…์ธ ๋ฅผ ์‰ฝ๊ณ  ๋น ๋ฅด๊ฒŒ ์ž‘์„ฑํ•ด ๋“œ๋ฆฝ๋‹ˆ๋‹ค.
    - ์Šคํ† ์–ด์—์„œ ๋ฌด๋ฃŒ ์บ์‹œ๋ฅผ ๊ณ„์ •๋ณ„๋กœ 1ํšŒ ๋ฐœ๊ธ‰ ๋ฐ›์„ ์ˆ˜ ์žˆ์Šต๋‹ˆ๋‹ค. ์ง€๊ธˆ ๋ฐ”๋กœ ์ฒดํ—˜ํ•ด ๋ณด์„ธ์š”!
    ์ด๋Ÿฐ ์ฃผ์ œ๋“ค์„ ์ž…๋ ฅํ•ด ๋ณด์„ธ์š”.
    - ์œ ์•„์—๊ฒŒ ์ ํ•ฉํ•œ ๋ฌธํ•™์ž‘ํ’ˆ์˜ ๊ธฐ์ค€๊ณผ ํŠน์„ฑ
    - ํ•œ๊ตญ์ธ์˜ ๊ฐ€์น˜๊ด€ ์ค‘์—์„œ ์ •์‹ ์  ๊ฐ€์น˜๊ด€์„ ์ด๋ฃจ๋Š” ๊ฒƒ๋“ค์„ ๋ฌธํ™”์  ๋ฌธ๋ฒ•์œผ๋กœ ์ •๋ฆฌํ•˜๊ณ , ํ˜„๋Œ€ํ•œ๊ตญ์‚ฌํšŒ์—์„œ ์ผ์–ด๋‚˜๋Š” ์‚ฌ๊ฑด๊ณผ ์‚ฌ๊ณ ๋ฅผ ๋น„๊ตํ•˜์—ฌ ์ž์‹ ์˜ ์˜๊ฒฌ์œผ๋กœ ๊ธฐ์ˆ ํ•˜์„ธ์š”
    - ์ž‘๋ณ„์ธ์‚ฌ ๋…ํ›„๊ฐ
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