ΠŸΠΎΠΌΠΎΡ‰ΡŒ Π² написании студСнчСских Ρ€Π°Π±ΠΎΡ‚
АнтистрСссовый сСрвис

ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½ΠΎΠ³ΠΎ супСрпродуцСнта Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ C (RACK1) с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ гСнСтичСской сСлСкции сайта ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции

Π”ΠΈΡΡΠ΅Ρ€Ρ‚Π°Ρ†ΠΈΡΠŸΠΎΠΌΠΎΡ‰ΡŒ Π² Π½Π°ΠΏΠΈΡΠ°Π½ΠΈΠΈΠ£Π·Π½Π°Ρ‚ΡŒ ΡΡ‚ΠΎΠΈΠΌΠΎΡΡ‚ΡŒΠΌΠΎΠ΅ΠΉ Ρ€Π°Π±ΠΎΡ‚Ρ‹

Π’Ρ‹Π²ΠΎΠ΄Ρ‹. ΠšΠΎΠΌΠΏΠ°Ρ€Ρ‚ΠΌΠ΅Π½Ρ‚Π°Π»ΠΈΠ·Π°Ρ†ΠΈΡ ΠΏΡ€ΠΎΠ΄ΡƒΡ†ΠΈΡ€ΡƒΠ΅ΠΌΠΎΠ³ΠΎ Π±Π΅Π»ΠΊΠ°. Π“Π»Π°Π²Π° I. Π Π΅ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Π½Ρ‚Π½Ρ‹Π΅ Π±Π΅Π»ΠΊΠΈ ΠΊΠ°ΠΊ инструмСнт для изучСния Π²Π½ΡƒΡ‚Ρ€ΠΈΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½ΠΎΠΉ сигнализации (Π½Π° ΠΏΡ€ΠΈΠΌΠ΅Ρ€Π΅ Π±Π΅Π»ΠΊΠ° RACK1).ΠžΠ±Π·ΠΎΡ€ Π»ΠΈΡ‚Π΅Ρ€Π°Ρ‚ΡƒΡ€Ρ‹. ΠŸΡ€ΠΈΠ½Ρ†ΠΈΠΏ ΠΈ Π΄ΠΈΠ·Π°ΠΉΠ½ систСмы гСнСтичСской сСлСкции. Амплификация Π”ΠΠš in vitro. Π₯ΠΈΠΌΠ΅Ρ€Π½Ρ‹Π΅ Π±Π΅Π»ΠΊΠΈ. Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅ Ρ€Π°Π½Π΄ΠΎΠΌΠΈΠ·ΠΎΠ²Π°Π½Π½ΠΎΠ³ΠΎ ΠΎΠ»ΠΈΠ³ΠΎΠ½ΡƒΠΊΠ»Π΅ΠΎΡ‚ΠΈΠ΄Π° Π² ΠΊΠΎΠ½ΡΡ‚Ρ€ΡƒΠΊΡ†ΠΈΡŽ pGCT2/pGCT. Π‘ΡƒΡ„Π΅Ρ€Ρ‹ ΠΈ Ρ€Π°ΡΡ‚Π²ΠΎΡ€Ρ‹. Π›Π°Π±ΠΎΡ€Π°Ρ‚ΠΎΡ€Π½ΠΎΠ΅ ΠΎΠ±ΠΎΡ€ΡƒΠ΄ΠΎΠ²Π°Π½ΠΈΠ΅. 81… Π§ΠΈΡ‚Π°Ρ‚ΡŒ Π΅Ρ‰Ρ‘ >

ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½ΠΎΠ³ΠΎ супСрпродуцСнта Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ C (RACK1) с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ гСнСтичСской сСлСкции сайта ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции (Ρ€Π΅Ρ„Π΅Ρ€Π°Ρ‚, курсовая, Π΄ΠΈΠΏΠ»ΠΎΠΌ, ΠΊΠΎΠ½Ρ‚Ρ€ΠΎΠ»ΡŒΠ½Π°Ρ)

Π‘ΠΎΠ΄Π΅Ρ€ΠΆΠ°Π½ΠΈΠ΅

  • Бписок сокращСний

Π“Π»Π°Π²Π° I. Π Π΅ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Π½Ρ‚Π½Ρ‹Π΅ Π±Π΅Π»ΠΊΠΈ ΠΊΠ°ΠΊ инструмСнт для изучСния Π²Π½ΡƒΡ‚Ρ€ΠΈΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½ΠΎΠΉ сигнализации (Π½Π° ΠΏΡ€ΠΈΠΌΠ΅Ρ€Π΅ Π±Π΅Π»ΠΊΠ° RACK1).ΠžΠ±Π·ΠΎΡ€ Π»ΠΈΡ‚Π΅Ρ€Π°Ρ‚ΡƒΡ€Ρ‹.

1.1. Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅.

1.2. Π‘Π΅Π»ΠΎΠΊ RACK1 — участник фосфоинозитидного ΠΏΡƒΡ‚ΠΈ ΠΏΠ΅Ρ€Π΅Π΄Π°Ρ‡ΠΈ сигнала. 10 Ѐосфоинозитидный ΠΏΡƒΡ‚ΡŒ ΠΏΠ΅Ρ€Π΅Π΄Π°Ρ‡ΠΈ сигнала. 10 ΠŸΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Π° Π‘ ΠΈ Π΅Π΅ Π²Π½ΡƒΡ‚Ρ€ΠΈΠΊΠ»Π΅Ρ‚ΠΎΡ‡Π½Ρ‹Π΅ Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Ρ‹. 11 RACK1 — Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€ Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ Π‘.

1.3. ЭкспрСссия Ρ‡ΡƒΠΆΠ΅Ρ€ΠΎΠ΄Π½Ρ‹Ρ… Π±Π΅Π»ΠΊΠΎΠ² Π² Π•.coli.

1.3.1. ГСнСтичСскиС элСмСнты ΡΠΊΡΠΏΡ€Π΅ΡΡΠΈΡ€ΡƒΡŽΡ‰Π΅Π³ΠΎ Π²Π΅ΠΊΡ‚ΠΎΡ€Π°. 17 ΠŸΡ€ΠΎΠΌΠΎΡ‚ΠΎΡ€Ρ‹. 17 Π Π΅ΡˆΠ°ΡŽΡ‰Π°Ρ Ρ€ΠΎΠ»ΡŒ ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ Π² ΡΡ„фСктивности процСсса трансляции. 20 ЗакономСрности ΠΏΠ΅Ρ€Π²ΠΈΡ‡Π½ΠΎΠΉ структуры области ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции. 22 ВлияниС Π²Ρ‚ΠΎΡ€ΠΈΡ‡Π½ΠΎΠΉ структуры RBS Π½Π° ΡΡ„Ρ„Π΅ΠΊΡ‚ΠΈΠ²Π½ΠΎΡΡ‚ΡŒ ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции. 26 ΠŸΡ€ΠΈΠΌΠ΅Π½Π΅Π½ΠΈΠ΅ ΡΠΈΠ»ΡŒΠ½Ρ‹Ρ… ΠΏΡ€ΠΈΡ€ΠΎΠ΄Π½Ρ‹Ρ… RBS Π² Π³Π΅Π½Π½ΠΎΠΈΠ½ΠΆΠ΅Π½Π΅Ρ€Π½Ρ‹Ρ… конструкциях. 28 УсилитСли трансляции. 29 Π’Π΅Ρ€ΠΌΠΈΠ½Π°Ρ‚ΠΎΡ€Ρ‹ трансляции. 32 Π‘Ρ‚Π°Π±ΠΈΠ»ΠΈΠ·Π°Ρ‚ΠΎΡ€Ρ‹ мРНК. 33 ИспользованиС ΠΊΠΎΠ΄ΠΎΠ½ΠΎΠ².

1.3.2. ΠšΠΎΠΌΠΏΠ°Ρ€Ρ‚ΠΌΠ΅Π½Ρ‚Π°Π»ΠΈΠ·Π°Ρ†ΠΈΡ ΠΏΡ€ΠΎΠ΄ΡƒΡ†ΠΈΡ€ΡƒΠ΅ΠΌΠΎΠ³ΠΎ Π±Π΅Π»ΠΊΠ°.

1.3.3. ЭкспрСссия ΠΏΡ€Π°Π²ΠΈΠ»ΡŒΠ½ΠΎ ΡΠ²ΠΎΡ€Π°Ρ‡ΠΈΠ²Π°ΡŽΡ‰ΠΈΡ…ΡΡ Π±Π΅Π»ΠΊΠΎΠ² Π² Π•. coli.

1.3.4. Π₯ΠΈΠΌΠ΅Ρ€Π½Ρ‹Π΅ Π±Π΅Π»ΠΊΠΈ.

1.4. Рандомизация ΠΈ Π³Π΅Π½Π΅Ρ‚ичСская сСлСкция ΠΊΠ°ΠΊ инструмСнт молСкулярной Π±ΠΈΠΎΠ»ΠΎΠ³ΠΈΠΈ.

Π“Π»Π°Π²Π° II. ΠŸΠΎΠ»ΡƒΡ‡Π΅Π½ΠΈΠ΅ Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½ΠΎΠ³ΠΎ супСрпродуцСнта — Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ Π‘ (RACK1) с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ гСнСтичСской сСлСкции сайта ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции. Π Π΅Π·ΡƒΠ»ΡŒΡ‚Π°Ρ‚Ρ‹ ΠΈ ΠΎΠ±ΡΡƒΠΆΠ΄Π΅Π½ΠΈΠ΅.

2.1. ΠŸΡ€ΠΈΠ½Ρ†ΠΈΠΏ ΠΈ Π΄ΠΈΠ·Π°ΠΉΠ½ систСмы гСнСтичСской сСлСкции.

2.2. ΠšΠΎΠ½ΡΡ‚Ρ€ΡƒΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΈ ΠΊΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ синтСтичСского Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚Π° COTR Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Π½ΠΎΠΌ Π²Π΅ΠΊΡ‚ΠΎΡ€Π΅ pGEM 1.

2.3. ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΎΠ΄ΠΈΡ€ΡƒΡŽΡ‰Π΅ΠΉ части Π³Π΅Π½Π° рСзистСнтности ΠΊ Ρ‚Π΅Ρ‚Ρ€Π°Ρ†ΠΈΠΊΠ»ΠΈΠ½Ρƒ Tetr Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Ρƒ pGEM-C2.

2.4. Π‘Ρ…Π΅ΠΌΠ° ввСдСния Ρ€Π°Π½Π΄ΠΎΠΌΠΈΠ·ΠΎΠ²Π°Π½Π½ΠΎΠ³ΠΎ Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚Π°.

2.5. ΠŸΡ€ΠΎΠ²Π΅Π΄Π΅Π½ΠΈΠ΅ гСнСтичСской сСлСкции Π² ΠΎΠ΄Π½ΠΎΡ†ΠΈΡΡ‚Ρ€ΠΎΠ½Π½ΠΎΠΉ систСмС.

2.6. ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠ”ΠΠš RACK1 Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Ρƒ pGCT2.

2.7. ВСстированиС ΠΏΠΎΠ»ΡƒΡ‡Π΅Π½Π½ΠΎΠ³ΠΎ сайта ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции для одноцистронной систСмы (RBS-Tet) Π² Π΄Π²ΡƒΡ†ΠΈΡΡ‚Ρ€ΠΎΠ½Π½ΠΎΠΉ конструкции, содСрТащСй гаск.

2.8. ΠšΠΎΠ½Ρ‚Ρ€ΠΎΠ»ΡŒΠ½Π°Ρ конструкция для экспрСссии RACK1 Π² Π΄Π²ΡƒΡ†ΠΈΡΡ‚Ρ€ΠΎΠ½Π½ΠΎΠΉ систСмС.

2.9. Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅ Ρ€Π°Π½Π΄ΠΎΠΌΠΈΠ·ΠΎΠ²Π°Π½Π½ΠΎΠ³ΠΎ Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚Π° Π² Π΄Π²ΡƒΡ†ΠΈΡΡ‚Ρ€ΠΎΠ½Π½ΡƒΡŽ ΠΊΠΎΠ½ΡΡ‚Ρ€ΡƒΠΊΡ†ΠΈΡŽ pGCT-RACK1.

2.10. Π˜ΡΠΊΡƒΡΡΡ‚Π²Π΅Π½Π½Ρ‹ΠΉ ΠΎΡ‚Π±ΠΎΡ€ ΠΈ Π°Π½Π°Π»ΠΈΠ· ΠΎΡ‚Π΄Π΅Π»ΡŒΠ½Ρ‹Ρ… ΠΊΠ»ΠΎΠ½ΠΎΠ².

2.11. ВСстированиС области ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции ΠΈΠ· ΠΊΠ»ΠΎΠ½Π°-супСрпродуцСнта Π² ΠΈΡΡ…ΠΎΠ΄Π½ΠΎΠΌ Π²Π΅ΠΊΡ‚ΠΎΡ€Π΅ pGCT2.

Π“Π»Π°Π²Π° III. ΠœΠ°Ρ‚Π΅Ρ€ΠΈΠ°Π»Ρ‹ ΠΈ ΠΌΠ΅Ρ‚ΠΎΠ΄Ρ‹.

3.1. Π Π΅Π°ΠΊΡ‚ΠΈΠ²Ρ‹.

3.2. Π€Π΅Ρ€ΠΌΠ΅Π½Ρ‚Ρ‹. 78 3.3 ΠœΠ°Ρ‚Π΅Ρ€ΠΈΠ°Π»Ρ‹.

3.4. ΠŸΡ€Π΅ΠΏΠ°Ρ€Π°Ρ‚Ρ‹.

3.5. ΠžΠ»ΠΈΠ³ΠΎΠ½ΡƒΠΊΠ»Π΅ΠΎΡ‚ΠΈΠ΄Ρ‹.

3.6. Π¨Ρ‚Π°ΠΌΠΌΡ‹ ΠΈ Π²Π΅ΠΊΡ‚ΠΎΡ€Ρ‹.

3.7. Π›Π°Π±ΠΎΡ€Π°Ρ‚ΠΎΡ€Π½ΠΎΠ΅ ΠΎΠ±ΠΎΡ€ΡƒΠ΄ΠΎΠ²Π°Π½ΠΈΠ΅. 81.

3.8. Π‘ΡƒΡ„Π΅Ρ€Ρ‹ ΠΈ Ρ€Π°ΡΡ‚Π²ΠΎΡ€Ρ‹.

3.9. ΠœΠ΅Ρ‚ΠΎΠ΄Ρ‹.

Π­Π»Π΅ΠΊΡ‚Ρ€ΠΎΡ„ΠΎΡ€Π΅Π· Π½ΡƒΠΊΠ»Π΅ΠΈΠ½ΠΎΠ²Ρ‹Ρ… кислот Π² Π½Π΅Π΄Π΅Π½Π°Ρ‚ΡƒΡ€ΠΈΡ€ΡƒΡŽΡ‰Π΅ΠΌ ΠŸΠΠΠ“.

Врансформация Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½Ρ‹Ρ… ΠΊΠ»Π΅Ρ‚ΠΎΠΊ ΠΈ ΠΎΡ‚Π±ΠΎΡ€ Ρ€Π΅ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Π½Ρ‚ΠΎΠ². 84 ΠŸΡ€Π΅ΠΏΠ°Ρ€Π°Ρ‚ΠΈΠ²Π½ΠΎΠ΅ Π²Ρ‹Π΄Π΅Π»Π΅Π½ΠΈΠ΅ ΠΈ ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Π½ΠΎΠΉ Π”ΠΠš ΠΌΠ΅Ρ‚ΠΎΠ΄ΠΎΠΌ Ρ‰Π΅Π»ΠΎΡ‡Π½ΠΎΠ³ΠΎ лизиса ΠΈ ΠΎΡ‡ΠΈΡΡ‚ΠΊΠ° Π½Π° ΠΈΠΎΠ½ΠΎΠΎΠ±ΠΌΠ΅Π½Π½Ρ‹Ρ… ΠΊΠΎΠ»ΠΎΠ½ΠΊΠ°Ρ… «Qiagen-tip 500».

ΠŸΡ€ΠΈΠ³ΠΎΡ‚ΠΎΠ²Π»Π΅Π½ΠΈΠ΅ Π»ΠΈΠ·Π°Ρ‚ΠΎΠ² ΠΊΠ»Π΅Ρ‚ΠΎΠΊ для Π±Π΅Π»ΠΊΠΎΠ²ΠΎΠ³ΠΎ элСктрофорСза.

Π‘Π΅Π»ΠΊΠΎΠ²Ρ‹ΠΉ элСктрофорСз.

Амплификация Π”ΠΠš in vitro.

ΠžΡ‡ΠΈΡΡ‚ΠΊΠ° ΠΏΡ€ΠΎΠ΄ΡƒΠΊΡ‚ΠΎΠ² Π°ΠΌΠΏΠ»ΠΈΡ„ΠΈΠΊΠ°Ρ†ΠΈΠΈ.

Π˜Π·Π²Π»Π΅Ρ‡Π΅Π½ΠΈΠ΅ Π”ΠΠš ΠΈΠ· Π°Π³Π°Ρ€ΠΎΠ·Π½ΠΎΠ³ΠΎ гСля.

Π˜ΠΌΠΌΡƒΠ½ΠΎΠ±Π»ΠΎΡ‚ΠΈΠ½Π³.

ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ синтСтичСского дуплСкса COTR Π² ΡΠΎΡΡ‚Π°Π²Π΅ ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Ρ‹ pGEMl. 90 ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΎΠ΄ΠΈΡ€ΡƒΡŽΡ‰Π΅ΠΉ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Π³Π΅Π½Π° устойчивости ΠΊ Ρ‚Π΅Ρ‚Ρ€Π°Ρ†ΠΈΠΊΠ»ΠΈΠ½Ρƒ Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Ρƒ pGEMC2. 91 ΠšΠ»ΠΎΠ½ΠΈΡ€ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΊΠΎΠ΄ΠΈΡ€ΡƒΡŽΡ‰Π΅ΠΉ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Π³Π΅Π½Π° Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ Π‘ — RACK1 Π² ΡΠΎΡΡ‚Π°Π²Π΅ ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Ρ‹ pGCT2. 92

Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅ Ρ€Π°Π½Π΄ΠΎΠΌΠΈΠ·ΠΎΠ²Π°Π½Π½ΠΎΠ³ΠΎ ΠΎΠ»ΠΈΠ³ΠΎΠ½ΡƒΠΊΠ»Π΅ΠΎΡ‚ΠΈΠ΄Π° Π² ΠΊΠΎΠ½ΡΡ‚Ρ€ΡƒΠΊΡ†ΠΈΡŽ pGCT2/pGCT

RACK1.

Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅ синтСтичСского дуплСкса RBS-Tet Π² Π²Π΅ΠΊΡ‚ΠΎΡ€ pGCT-RACKl.

Π’Π²Π΅Π΄Π΅Π½ΠΈΠ΅ синтСтичСского дуплСкса SD Π² ΠΊΠΎΠ½ΡΡ‚Ρ€ΡƒΠΊΡ†ΠΈΡŽ pGCT-RACKl.

Π˜ΡΠΊΡƒΡΡΡ‚Π²Π΅Π½Π½Ρ‹ΠΉ ΠΎΡ‚Π±ΠΎΡ€.

Π’Ρ‹Π²ΠΎΠ΄Ρ‹.

Π‘ΠΎΠ²Π΅Ρ€ΡˆΠ΅Π½ΡΡ‚Π²ΠΎΠ²Π°Π½ΠΈΠ΅ ΠΌΠ΅Ρ‚ΠΎΠ΄ΠΎΠ»ΠΎΠ³ΠΈΠΈ Ρ€Π΅ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Π½Ρ‚Π½Ρ‹Ρ… Π”ΠΠš ΠΏΠΎΡ€Π°Π·ΠΈΡ‚Π΅Π»ΡŒΠ½ΠΎ Ρ€Π°ΡΡˆΠΈΡ€ΠΈΠ»ΠΎ возмоТности Π½Π°ΠΏΡ€Π°Π²Π»Π΅Π½Π½ΠΎΠ³ΠΎ биосинтСза Π±Π΅Π»ΠΊΠ°. Π‘Ρ€Π΅Π΄ΠΈ Ρ€Π°Π·Π»ΠΈΡ‡Π½Ρ‹Ρ… систСм экспрСссии гСнСтичСских Π΄Π΅Ρ‚Π΅Ρ€ΠΌΠΈΠ½Π°Π½Ρ‚ Π²Π°ΠΆΠ½ΠΎΠ΅ мСсто Π·Π°Π½ΠΈΠΌΠ°ΡŽΡ‚ прокариотичСскиС систСмы. Π‘Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½Ρ‹Π΅ супСрпродуцСнты ΠΏΠΎΠ·Π²ΠΎΠ»ΡΡŽΡ‚ ΠΏΠΎΠ»ΡƒΡ‡Π°Ρ‚ΡŒ Π·Π½Π°Ρ‡ΠΈΡ‚Π΅Π»ΡŒΠ½Ρ‹Π΅ количСства высокоочищСнных Ρ€Π΅ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Π½Ρ‚Π½Ρ‹Ρ… Π±Π΅Π»ΠΊΠΎΠ² для использования Π² ΠΌΠ΅Π΄ΠΈΠΊΠΎ-биологичСских ΠΈ ΡΠ΅Π»ΡŒΡΠΊΠΎΡ…озяйствСнных цСлях.

Для экспрСссии Π² Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½ΠΎΠΉ систСмС Π±Π΅Π·ΠΈΠ½Ρ€ΠΎΠ½Π½Ρ‹ΠΉ Π³Π΅Π½ (ΠΊΠ”ΠΠš) ΠΊΠ»ΠΎΠ½ΠΈΡ€ΡƒΡŽΡ‚ Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Π½Ρ‹ΠΉ Π²Π΅ΠΊΡ‚ΠΎΡ€, содСрТащий структурныС элСмСнты, Π½Π΅ΠΎΠ±Ρ…ΠΎΠ΄ΠΈΠΌΡ‹Π΅ для Π΅Π³ΠΎ транскрипции ΠΈ Ρ‚рансляции. НаиболСС эффСктивная ΠΈ ΡˆΠΈΡ€ΠΎΠΊΠΎ ΠΈΡΠΏΠΎΠ»ΡŒΠ·ΡƒΠ΅ΠΌΠ°Ρ систСма транскрипции (систСма Π‘Ρ‚ΡƒΠ΄ΠΈΠ΅Ρ€Π°) Π²ΠΊΠ»ΡŽΡ‡Π°Π΅Ρ‚ ΠΏΡ€ΠΎΠΌΠΎΡ‚ΠΎΡ€ ΠΈΠ· Ρ„Π°Π³Π° Π’7 Π² ΡΠΎΡ‡Π΅Ρ‚Π°Π½ΠΈΠΈ с Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½Ρ‹ΠΌΠΈ ΡˆΡ‚Π°ΠΌΠΌΠ°ΠΌΠΈ, нСсущими ΠΈΠ½Π΄ΡƒΡ†ΠΈΠ±Π΅Π»ΡŒΠ½Ρ‹ΠΉ Π³Π΅Π½ Π’7-РНК-ΠΏΠΎΠ»ΠΈΠΌΠ΅Ρ€Π°Π·Ρ‹ Π² ΡΠ²ΠΎΠ΅ΠΌ Π³Π΅Π½ΠΎΠΌΠ΅ (Studier & Moffat, 1986). Π­Ρ‚Π° систСма обСспСчиваСт ΠΌΠ°ΡΡΠΈΠ²Π½ΡƒΡŽ Ρ‚Ρ€Π°Π½ΡΠΊΡ€ΠΈΠΏΡ†ΠΈΡŽ, ΡΡ„Ρ„Π΅ΠΊΡ‚ΠΈΠ²Π½ΠΎΡΡ‚ΡŒ ΠΊΠΎΡ‚ΠΎΡ€ΠΎΠΉ практичСски Π½Π΅ Π·Π°Π²ΠΈΡΠΈΡ‚ ΠΎΡ‚ ΡΡ‚Ρ€ΡƒΠΊΡ‚ΡƒΡ€Ρ‹ Ρ†Π΅Π»Π΅Π²ΠΎΠ³ΠΎ Π³Π΅Π½Π°. Однако, Π²Ρ‹Ρ…ΠΎΠ΄ Π±Π΅Π»ΠΊΠ° Π² ΠΎΠ΄Π½ΠΎΠΉ ΠΈ Ρ‚ΠΎΠΉ ΠΆΠ΅ систСмС экспрСссии ΠΌΠΎΠΆΠ΅Ρ‚ сильно Π²Π°Ρ€ΡŒΠΈΡ€ΠΎΠ²Π°Ρ‚ΡŒ ΠΎΡ‚ Π³Π΅Π½Π° ΠΊ Π³Π΅Π½Ρƒ. Π­Ρ‚ΠΎΡ‚ Ρ„Π°ΠΊΡ‚ ΠΎΠ±ΡŠΡΡΠ½ΡΠ΅Ρ‚ΡΡ Ρ‚Π΅ΠΌ, Ρ‡Ρ‚ΠΎ Π»ΠΈΠΌΠΈΡ‚ΠΈΡ€ΡƒΡŽΡ‰Π΅ΠΉ Π² ΡΡ‚ΠΈΡ… условиях являСтся ΡΠ»Π΅Π΄ΡƒΡŽΡ‰Π°Ρ стадии биосинтСза Π±Π΅Π»ΠΊΠ° — трансляция, Π² ΠΎΡΠΎΠ±Π΅Π½Π½ΠΎΡΡ‚ΠΈ Π΅Π΅ ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΡ. Π­Ρ‚Π° стадия являСтся Π½Π°ΠΈΠΌΠ΅Π½Π΅Π΅ прСдсказуСмой, Ρ‚Π°ΠΊ ΠΊΠ°ΠΊ Π΅Π΅ ΡΡ„Ρ„Π΅ΠΊΡ‚ΠΈΠ²Π½ΠΎΡΡ‚ΡŒ сильно зависит ΠΎΡ‚ ΡΡ‚Ρ€ΡƒΠΊΡ‚ΡƒΡ€Ρ‹ Ρ†Π΅Π»Π΅Π²ΠΎΠ³ΠΎ Π³Π΅Π½Π°, которая Π² Π±ΠΎΠ»ΡŒΡˆΠΎΠΉ стСпСни опрСдСляСт Π²Ρ‚ΠΎΡ€ΠΈΡ‡Π½ΡƒΡŽ структуру участка связывания рибосом.

Π’ ΠΈΠ΄Π΅Π°Π»Π΅ сайт связывания рибосом Π½ΡƒΠΆΠ½ΠΎ Π±Ρ‹Π»ΠΎ Π±Ρ‹ ΠΏΠΎΠ΄Π±ΠΈΡ€Π°Ρ‚ΡŒ ΡΠΏΠ΅Ρ†ΠΈΠ°Π»ΡŒΠ½ΠΎ для ΠΊΠ°ΠΆΠ΄ΠΎΠ³ΠΎ Ρ†Π΅Π»Π΅Π²ΠΎΠ³ΠΎ Π³Π΅Π½Π°, ΠΎΠ΄Π½Π°ΠΊΠΎ Ρ€Π°Ρ†ΠΈΠΎΠ½Π°Π»ΡŒΠ½ΠΎΠΉ основы для Ρ‚Π°ΠΊΠΎΠ³ΠΎ ΠΏΠΎΠ΄Π±ΠΎΡ€Π° Π½Π΅ ΡΡƒΡ‰Π΅ΡΡ‚Π²ΡƒΠ΅Ρ‚, Ρ‚Π°ΠΊ Ρ‡Ρ‚ΠΎ ΠΎΠ±Ρ‹Ρ‡Π½ΠΎ это дСлаСтся ΠΌΠ΅Ρ‚ΠΎΠ΄ΠΎΠΌ ΠΏΡ€ΠΎΠ± ΠΈ ΠΎΡˆΠΈΠ±ΠΎΠΊ. Π’ Π½Π°ΡˆΠ΅ΠΉ Ρ€Π°Π±ΠΎΡ‚Π΅, вмСсто Ρ€ΡƒΡ‚ΠΈΠ½Π½ΠΎΠ³ΠΎ ΠΏΠ΅Ρ€Π΅Π±ΠΎΡ€Π° Π²Π°Ρ€ΠΈΠ°Π½Ρ‚ΠΎΠ² Ρ€Π°Π·Π»ΠΈΡ‡Π½Ρ‹Ρ… ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚Π΅ΠΉ, ΠΌΡ‹ Ρ€Π°Π·Ρ€Π°Π±ΠΎΡ‚Π°Π»ΠΈ ΠΊΠΎΠΌΠ±ΠΈΠ½Π°Ρ‚ΠΎΡ€Π½Ρ‹ΠΉ ΠΏΠΎΠ΄Ρ…ΠΎΠ΄ ΠΏΡ€ΠΈ ΠΊΠΎΡ‚ΠΎΡ€ΠΎΠΌ Π½ΡƒΠΆΠ½Ρ‹Π΅ ΠΏΠΎΡΠ»Π΅Π΄ΠΎΠ²Π°Ρ‚Π΅Π»ΡŒΠ½ΠΎΡΡ‚ΠΈ Π²Ρ‹Π±ΠΈΡ€Π°ΡŽΡ‚ΡΡ автоматичСски Π² Ρ€Π΅Π·ΡƒΠ»ΡŒΡ‚Π°Ρ‚Π΅ гСнСтичСской сСлСкции Π² ΡΠΏΠ΅Ρ†ΠΈΠ°Π»ΡŒΠ½ΠΎΠΉ систСмС.

Π­Ρ‚ΠΎΡ‚ ΠΏΠΎΠ΄Ρ…ΠΎΠ΄ позволяСт ΠΏΠΎΠ΄ΠΎΠ±Ρ€Π°Ρ‚ΡŒ эффСктивный сайт ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции для любого Ρ‡ΡƒΠΆΠ΅Ρ€ΠΎΠ΄Π½ΠΎΠ³ΠΎ Π³Π΅Π½Π° Π² ΠΎΠ΄Π½ΠΎΠΌ экспСримСнтС. Π­Ρ‚Π° систСма Π±Ρ‹Π»Π° Π½Π°ΠΌΠΈ использована для создания супСрпродуцСнта Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ Π‘ — RACK1.

Π’Ρ‹Π²ΠΎΠ΄Ρ‹:

1. Π‘ΠΎΠ·Π΄Π°Π½Π° гСнСтичСская систСма, Π² ΠΊΠΎΡ‚ΠΎΡ€ΠΎΠΉ с ΠΏΠΎΠΌΠΎΡ‰ΡŒΡŽ Π½Π°ΠΏΡ€Π°Π²Π»Π΅Π½Π½ΠΎΠΉ ΡΠ²ΠΎΠ»ΡŽΡ†ΠΈΠΈ ΠΌΠΎΠΆΠ½ΠΎ ΠΎΠΏΡ‚ΠΈΠΌΠΈΠ·ΠΈΡ€ΠΎΠ²Π°Ρ‚ΡŒ сайт ΠΈΠ½ΠΈΡ†ΠΈΠ°Ρ†ΠΈΠΈ трансляции Ρ‡ΡƒΠΆΠ΅Ρ€ΠΎΠ΄Π½ΠΎΠ³ΠΎ Π³Π΅Π½Π° для экспрСссии Π² Π±Π°ΠΊΡ‚Π΅Ρ€ΠΈΠ°Π»ΡŒΠ½ΠΎΠΉ систСмС.

2. Π Π°Π·Ρ€Π°Π±ΠΎΡ‚Π°Π½ Π½ΠΎΠ²Ρ‹ΠΉ ΠΌΠ΅Ρ‚ΠΎΠ΄ ввСдСния ΠΊΠΎΡ€ΠΎΡ‚ΠΊΠΎΠ³ΠΎ Ρ€Π°Π½Π΄ΠΎΠΌΠΈΠ·ΠΎΠ²Π°Π½Π½ΠΎΠ³ΠΎ Ρ„Ρ€Π°Π³ΠΌΠ΅Π½Ρ‚Π° Π² ΠΏΠ»Π°Π·ΠΌΠΈΠ΄Π½Ρ‹ΠΉ Π²Π΅ΠΊΡ‚ΠΎΡ€.

3. Π‘ ΠΈΡΠΏΠΎΠ»ΡŒΠ·ΠΎΠ²Π°Π½ΠΈΠ΅ΠΌ Ρ€Π°Π·Ρ€Π°Π±ΠΎΡ‚Π°Π½Π½ΠΎΠΉ систСмы ΠΏΠΎΠ»ΡƒΡ‡Π΅Π½ супСрпродуцСнт Ρ€Π΅Ρ†Π΅ΠΏΡ‚ΠΎΡ€Π° Π°ΠΊΡ‚ΠΈΠ²ΠΈΡ€ΠΎΠ²Π°Π½Π½ΠΎΠΉ ΠΏΡ€ΠΎΡ‚Π΅ΠΈΠ½ΠΊΠΈΠ½Π°Π·Ρ‹ Π‘ (RACK1) Ρ‡Π΅Π»ΠΎΠ²Π΅ΠΊΠ°.

ΠŸΠΎΠΊΠ°Π·Π°Ρ‚ΡŒ вСсь тСкст

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